Jutta K. Preiksaitis

Canadian Society of Transplantation Consensus Workshop on Cytomegalovirus Management in Solid Organ Transplantation Final Report

The Canadian Society of Transplantation sponsored a Cytomegalovirus (CMV) Consensus Working Group that met on March 19, 2003. The objectives of this group were to determine the current burden of CMV‐associated disease in the setting of solid organ transplantation in Canada, make recommendations regarding optimal strategies for the diagnosis, treatment and prevention of CMV infection and disease, highlight gaps in knowledge and outline priorities for research and other initiatives that might further reduce the burden of CMV‐associated effects in this setting. This report summarizes the recommendations of the working group including ratings of the strength of evidence supporting the recommendations.

Interlaboratory Comparison of Cytomegalovirus Viral Load Assays

To assess interlaboratory variability in qualitative and quantitative cytomegalovirus (CMV) viral load (VL) testing, we distributed a panel of samples to 33 laboratories in the USA, Canada and Europe who performed testing using commercial reagents (n = 17) or laboratory‐developed assays (n = 18). The panel included two negatives, seven samples constructed from purified CMV nucleocapsids in plasma (2.0–6.0 log10 copies/mL) and three clinical plasma samples. Interlaboratory variation was observed in both actual (range, 2.0–4.0 log10 copies/mL) and self‐reported lower limits of detection (range, 1.0–4.0 log10 copies/mL). Variation observed in reported results for individual samples ranged from 2.0 log10 (minimum) to 4.3 log10 (maximum). Variation was greatest at low VLs. Assuming ± 0.5 log10 relative to the expected result represents an acceptable result, 57.6% of results fell within this range. Use of commercially available reagents and procedures was associated with less variability compared with laboratory‐developed assays. Interlaboratory variability on replicate samples was significantly greater than intralaboratory variability (p < 0.0001). The significant interlaboratory variability in CMV VL observed may be impacting patient care and limiting interinstitutional comparisons. The creation of an international reference standard for CMV VL assay calibration would be an important step in quality improvement of this laboratory tool.

Herpes Zoster Infection Following Solid Organ Transplantation: Incidence, Risk Factors and Outcomes in the Current Immunosuppressive Era

Herpes zoster (HZ) infection is a frequent and serious complication of organ transplantation that has not been examined in the current era of immunosuppression.

Cell‐Mediated Immunity to Predict Cytomegalovirus Disease in High‐Risk Solid Organ Transplant Recipients

Late‐onset cytomegalovirus (CMV) disease commonly occurs after discontinuation of antiviral prophylaxis. We determined the utility of testing CD8+ T‐cell response against CMV as a predictor of late‐onset CMV disease after a standard course of antiviral prophylaxis. Transplant patients at high‐risk for CMV disease were enrolled. CD8+ T‐cell‐mediated immunity (CMI) was tested using the QuantiFERON‐CMV assay at baseline, 1, 2 and 3 months posttransplant by measurement of interferon‐γ response to whole blood stimulation with a 21‐peptide pool. The primary outcome was the ability of CMI testing to predict CMV disease in the first 6 months posttransplant. There were 108 evaluable patients (D+/R+ n = 39; D‐/R+ n = 34; D+/R‐ n = 35) of whom 18 (16.7%) developed symptomatic CMV disease. At the end of prophylaxis, CMI was detectable in 38/108 (35.2%) patients (cutoff 0.1 IU/mL interferon‐γ). CMV disease occurred in 2/38 (5.3%) patients with a detectable interferon‐γ response versus 16/70 (22.9%) patients with a negative response; p = 0.038. In the subgroup of D+/R‐ patients, CMV disease occurred in 1/10 (10.0%) patients with a detectable interferon‐γ response (cutoff 0.1 IU/mL) versus 10/25 (40.0%) patients with a negative CMI, p = 0.12. Monitoring of CMI may be useful for predicting late‐onset CMV disease.

Epstein-barr virus and posttransplant lymphoproliferative disorder in solid organ transplant recipients.

Posttransplant lymphoproliferative disorder (PTLD) is recognized as potentially one of the most devastating complications of organ transplantation. Epstein-Barr virus (EBV) is associated with the majority of PTLD cases. The entity referred to as EBV-associated PTLD encompasses a wide spectrum of clinical conditions characterized by lymphoproliferation after transplantation. These syndromes range from uncomplicated infectious mononucleosis to true malignancies (1–3). Disease may be nodal or extranodal, localized, often in the allograft, or widely disseminated. Patients may be symptomatic or asymptomatic. PTLD may resemble a self-limited infection or be indistinguishable from non-Hodgkin’s lymphoma. Lesions may be localized and progress slowly or the patient may present with a fulminant multisystem sepsis-like syndrome.

Interlaboratory comparison of epstein-barr virus viral load assays.

To assess interlaboratory variability in qualitative and quantitative Epstein‐Barr virus (EBV) viral load (VL) testing, we distributed a panel of samples to 28 laboratories in the USA, Canada and Europe who performed testing using commercially available reagents (n = 12) or laboratory‐developed assays (n = 18). The panel included two negatives, seven constructed samples using Namalwa and Molt‐3 cell lines diluted in plasma (1.30–5.30 log10 copies/mL) and three clinical plasma samples. Significant interlaboratory variation was observed for both actual (range 1.30–4.30 log10 copies/mL) and self‐reported (range, 1.70–3.30 log10 copies/mL) lower limits of detection. The variation observed in reported results on individual samples ranged from 2.28 log10 (minimum) to 4.14 log10 (maximum). Variation was independent of dynamic range and use of commercial versus laboratory‐developed assays. Overall, only 47.0% of all results fell within acceptable standards of variation: defined as the expected result ± 0.50 log10. Interlaboratory variability on replicate samples was significantly greater than intralaboratory variability (p < 0.0001). Kinetics of change in VL appears more relevant than absolute values and clinicians should understand the uncertainty associated with absolute VL values at their institutions. The creation of an international reference standard for EBV VL assay calibration would be an initial important step in quality improvement of this laboratory tool.

Nucleic acid testing (NAT) of organ donors: is the 'best' test the right test? A consensus conference report.

Nucleic acid testing (NAT) for HIV, HBV and HCV shortens the time between infection and detection by available testing. A group of experts was selected to develop recommendations for the use of NAT in the HIV/HBV/HCV screening of potential organ donors. The rapid turnaround times needed for donor testing and the risk of death while awaiting transplantation make organ donor screening different from screening blood‐or tissue donors. In donors with no identified risk factors, there is insufficient evidence to recommend routine NAT, as the benefits of NAT may not outweigh the disadvantages of NAT especially when false‐positive results can lead to loss of donor organs. For donors with identified behavioral risk factors, NAT should be considered to reduce the risk of transmission and increase organ utilization. Informed consent balancing the risks of donor‐derived infection against the risk of remaining on the waiting list should be obtained at the time of candidate listing and again at the time of organ offer. In conclusion, there is insufficient evidence to recommend universal prospective screening of organ donors for HIV, HCV and HBV using current NAT platforms. Further study of viral screening modalities may reduce disease transmission risk without excessive donor loss.

Comparison of Quantiferon‐TB Gold With Tuberculin Skin Test for Detecting Latent Tuberculosis Infection Prior to Liver Transplantation

Screening for latent tuberculosis infection (LTBI) is recommended prior to organ transplantation. The Quantiferon‐TB Gold assay (QFT‐G) may be more accurate than the tuberculin skin test (TST) in the detection of LTBI. We prospectively compared the results of QFT‐G to TST in patients with chronic liver disease awaiting transplantation. Patients were screened for LTBI with both the QFT‐G test and a TST. Concordance between test results and predictors of a discordant result were determined. Of the 153 evaluable patients, 37 (24.2%) had a positive TST and 34 (22.2%) had a positive QFT‐G. Overall agreement between tests was 85.1% (κ= 0.60, p < 0.0001). Discordant test results were seen in 12 TST positive/QFT‐G negative patients and in 9 TST negative/QFT‐G positive patients. Prior BCG vaccination was not associated with discordant test results. Twelve patients (7.8%), all with a negative TST, had an indeterminate result of the QFT‐G and this was more likely in patients with a low lymphocyte count (p = 0.01) and a high MELD score (p = 0.001). In patients awaiting liver transplantation, both the TST and QFT‐G were comparable for the diagnosis of LTBI with reasonable concordance between tests. Indeterminate QFT‐G result was more likely in those with more advanced liver disease.

A Trial of Valganciclovir Prophylaxis for Cytomegalovirus Prevention in Lung Transplant Recipients

Cytomegalovirus (CMV) infection is common after lung transplantation. We performed a prospective trial of valganciclovir prophylaxis in lung recipients with outcomes compared to matched historical controls. The valganciclovir group (n = 40) (including D+/R– and R+ patients) was prospectively enrolled, and received oral valganciclovir 900 mg once daily for 12 weeks. Historical controls (n = 40) received 12 weeks of daily intravenous ganciclovir if D+/R– or 12 weeks of oral ganciclovir if R+. CMV viral load testing was done at two‐week intervals until 6 months posttransplant. Baseline demographics and immunosuppression were comparable in the two groups. The incidence of CMV viremia was 16/40 (40.0%) in the valganciclovir arm versus 18/40 (45%) in the ganciclovir arm (p = NS). The incidence of symptomatic CMV disease was 8/40 (20%) versus 7/40 (17.5%), respectively (p = NS). In both groups viremia, while on prophylaxis, was uncommon (valganciclovir: 0/40 and ganciclovir: 2/40). Peak viral load and time to viremia were similar in the two arms. High rates of viremia and symptomatic disease occurred in the D+/R– patients after discontinuation of prophylaxis. Genotypic CMV sequence analysis demonstrated low rates of ganciclovir resistance in both groups. Valganciclovir prophylaxis had similar efficacy to either intravenous ganciclovir (D+/R– patients), or oral ganciclovir (R+ patients) in lung recipients.

Comparison of LightCycler-Based PCR, COBAS Amplicor CMV Monitor, and pp65 Antigenemia Assays for Quantitative Measurement of Cytomegalovirus Viral Load in Peripheral Blood Specimens from Patients after Solid Organ Transplantation

ABSTRACT In order to evaluate the LightCycler-based PCR (LC-PCR) as a diagnostic assay technique, a classical pp65 antigenemia assay and the commercially available COBAS Amplicor CMV Monitor (CACM) assay were compared to the LC-PCR assay for the detection and quantitation of cytomegalovirus (CMV) load in 404 parallel specimens of peripheral blood from 66 patients after solid organ transplantation. A good correlation existed among these three assays (r ≅ 0.6, P < 0.0001). The LC-PCR assay was the most sensitive (54% of specimens positive) compared to the CACM (48.6%) and the pp65 antigenemia (26%) assays. The LC-PCR assay detected all samples found positive by using both the CMV pp65 antigenemia assay and the CACM assay. The LC-PCR also had the widest dynamic range (from 250 to 107 DNA copies/ml of plasma). No cross-reactions were found among CMV and Epstein-Barr virus, varicella-zoster virus, or herpes simplex virus in the LC-PCR by using amplification with specifically designed primer pairs. Precision, expressed as the coefficient of variation, was <3% with standard DNA from cell cultures and between 6.55 and 14.1% with clinical specimens in repeat LC-PCR runs. One run of the LC-PCR took half of the time required for the semiautomated CACM procedure. Because of its sensitivity, specificity, cost-effectiveness, and simplicity, the LC-PCR assay could replace the pp65 antigenemia and the CACM assays as the preferred technique for the surveillance, diagnosis, and monitoring of response of CMV diseases in high-risk populations.