Real Time PCR and Serological Assays for diagnosis and monitoring of Human Cytomegalovirus Infection in Immunocompormised Pediatric Cancer Patient

Abstract

Background. Human cytomegalovirus (HCMV) is a major threat to pediatric leukemia patients.We performed this study to evaluate different detection methods for detection HCMV infection and determine virologic cut-off value associated with risk of CMV disease for starting pre-emptive therapy

Patients&Methods. Blood of fifty febrile leukemic children and 30 apparently healthy control was subjected for detection of HCMV in serum by serologic (IgM and IgG) assay and in both leukocytes and serum by molecular (qualitative PCR and quantitative PCR) assays and the results were compared.

Results. Positive rate of HCMV IgM was lower in leukemic children( 11/50 (22%) than control 13/30 (43%), P=0.044). But positivity of IgG did not show any difference between both groups (P=1.00). The highest positive results was detected in serum by qualitative PCR assay in 27/50 (54%) for leukemia and 14/30 (46.7%) for control children. Considering qualitative PCR assay is the golden standard assay for detection of active HCMV infection, sensitivity of CMV IgM and quantitative assay was lower in leukemic patients compared to control (40% and 70% respectively for leukemic patients vs 85.7% and 100% respectively for control group). In contrast sensitivity of high titer IgG was higher in leukemic patients than control (80% vs 50% respectively). The best DNAemia cut off value at 3.5x106 could identify leukemic patients at risk of CMV disease based on absolut lymphocytic count (P=0.037).

Conclusion. Qualitative PCR assay in serum was the most sensitive assay for detection presence of HCMV DNA among leukemic children followed by in house quantitative PCR assay. The best DNAemia cutoff value by quantitative PCR assay was at 3.52x106 copies/ml can discriminate patients with low and high absolute lymphocytic count and can be used for starting pre-emptive therapy. Further larger sample size analysis using serial sampling is required to confirm results.

Keywords: Human cytomegalovirus(HCMV), IgM immunoglobulin M, IgG immunoglobulin G, DNAemia,Leukemia,Cutoff, quantitative PCR assay.