Evaluation of the AMPLICOR Cytomegalovirus Test with Specimens from Human Immunodeficiency Virus-Infected Subjects

Abstract

The AMPLICOR cytomegalovirus (CMV) test, a new qualitative assay for the detection of CMV DNA in plasma, was compared to conventional methods and quantitative PCR (Q-PCR) assays by using leukocytes and plasma from 179 blood samples from subjects with AIDS. For the diagnosis of CMV disease, cell-based assays such as a Q-PCR with polymorphonuclear leukocytes (Q-PCR-PMNL) and a pp65 antigenemia assay had the highest sensitivities but suffered from a lack of specificity. The best agreement between the results of the Q-PCR-PMNL assay and those of the AMPLICOR test was found when a threshold diagnostic value of 690 copies per 105 cells was selected for the Q-PCR-PMNL assay. In that context, the AMPLICOR CMV test had a sensitivity of 96.4% and a specificity of 95.3% when results were compared to results of the cell-based PCR assay. This threshold was close to the one described as associated with the best sensitivity and specificity for the diagnosis of CMV disease in a recently published study (4). Blood samples that tested positive by the Q-PCR-PMNL assay but negative by the AMPLICOR CMV test were associated with viral loads (mean, 785 copies, median, 96 copies per 105leukocytes) lower than the viral loads of blood samples that tested positive by both assays (mean, 21,452 copies; median, 9,784 copies per 105 leukocytes) (P = 0.003). The AMPLICOR CMV test gave positive results at least 48 days before the development of symptomatic CMV disease in a longitudinal analysis of a limited subset of patients (n = 6) from whom sequential specimens were available for testing. In conclusion, the AMPLICOR CMV test is a very convenient assay combining rapidity, simplicity, and the possibility of batch testing. A positive result by this test seems particularly important since this implies, in most instances, the presence or the imminence of CMV disease, although a negative test result does not rule out disease.

Evaluation of New Quantitative Assays for Diagnosis and Monitoring of Cytomegalovirus Disease in Human Immunodeficiency Virus-Positive Patients

“…We observed few discordant results, and in most cases, when the results of two assays did not agree, the positive one was weakly positive while the other one was negative. Comparing a homemade PCR with PBLs and P-AMP with plasma, Boivin et al (5) also found that discordant results were associated with low viral loads. In addition, when we observed discordant results, the negative assay has often been positive previously and/or became positive later during the course of the CMV infection.…”

Review article: acute severe ulcerative colitis – evidence‐based consensus statements

“…Only two of eight studies on CMV infection in IBD demonstrated concordance between histology/IHC and tissue PCR . Whole blood leucocyte DNA PCR has diagnostic sensitivity of 65–100% and specificity of 40–92% . Colonic or blood CMV DNA may confirm CMV colitis but are not a prerequisite.…”

False-Positive Results of Plasma PCR for Cytomegalovirus DNA due to Delayed Sample Preparation

“…If cell turnover was a major mechanism that elicits CMV plasma DNAemia, this would suggest that in patients with CMV leukocyte DNAemia long-term storage of whole blood samples before separation may cause release of viral DNA into the plasma fraction, thus distorting PCR results. This question is of great importance not only in the clinical setting but also for test reproducibility and standardization, especially since commercial kits for CMV PCR of plasma samples are available (3,10). Some investigators argue that contamination of freshly prepared plasma fractions with cellular DNA can be overcome by ultrafiltration, whereas others have reported that PCR for cellular target sequences is regularly positive even with ultrafiltered samples (6,20,21).…”