Clinical evaluation of a quantitative real time polymerase chain reaction assay for diagnosis of primary Epstein-Barr virus infection in children

被引:47
作者
Pitetti, RD [1 ]
Laus, S
Wadowsky, RM
机构
[1] Univ Pittsburgh, Childrens Hosp Pittsburgh, Sch Med, Div Pediat Emergency Med,Dept Pediat, Pittsburgh, PA 15213 USA
[2] Univ Pittsburgh, Childrens Hosp Pittsburgh, Sch Med,Div Pediat Pathol, Dept Pathol, Pittsburgh, PA 15213 USA
[3] Univ Pittsburgh, Childrens Hosp Pittsburgh, Sch Med,Div Pediat Pathol, Dept Infect Dis & Microbiol, Pittsburgh, PA 15213 USA
[4] Childrens Hosp Pittsburgh, Pediat Mol Microbiol Lab, Pittsburgh, PA 15213 USA
[5] Univ Pittsburgh, Sch Med, Grad Sch Publ Hlth, Pittsburgh, PA USA
关键词
Epstein-Barr virus; infectious mononucleosis; polymerase chain reaction;
D O I
10.1097/01.inf.0000078157.90639.96
中图分类号
R392 [医学免疫学]; Q939.91 [免疫学];
学科分类号
100102 ;
摘要
Background. Epstein-Barr virus (EBV) infectious mononucleosis is often diagnosed based on characteristic clinical features and either a positive heterophil antibody test or serology, both of which can be unreliable in young children. Real time quantitative PCR assays that measure EBV DNA load in serum or plasma are highly sensitive in young children, but serum and plasma contain inhibitors of PCR which must be removed by DNA extraction techniques. A real time TaqMan PCR assay was designed and evaluated for simultaneously measuring EBV DNA load and validating the removal of PCR inhibitors from serum samples. Methods. A serum sample was available from patients classified serologically as primary EBV infection (n = 28), EBV-seronegative (n = 25) and EBV-seropositive (n = 26). Patients were classified as having EBV infectious mononucleosis if they had specified clinical findings and greater than or equal to10% atypical lymphocytes in peripheral blood or had a positive Monospot test result. DNA was purified by a spin column method and tested in PCR reactions with primers for EBV DNA polymerase gene and internal control targets. Amplification of the two PCR products was measured in real time with separate TaqMan DNA probes labeled with various fluorescent reporters. Results. The mean age of study patients was 9 years, 4 months. Twenty-one (75%) of the patients in the primary EBV infection group, one (4%) of the seronegatives and none of the seropositives had detectable EBV DNA. Within the primary infection group, those with detectable virus were more likely than those without detectable virus to have evidence of lymphadenopathy (14 of 16 vs. I of 5; P = 0.011), higher mean atypical (11.7 vs. 0.9%; P = 0.002) and absolute atypical (1.5 vs. 0.1 X 10(9)/l; P = 0.004) lymphocyte count, higher mean absolute lymphocyte count (4.7 vs. 2.3 X 10(9)/l; P = 0.026) and higher mean aspartate aminotransferase value (119.8 vs. 37.3 IU/l; P = 0.036). Ten patients, all in the primary infection group, had EBV infectious mononucleosis, and all had positive PCR results. No sample contained PCR inhibitors. Conclusions. A real time TaqMan PCR assay allows rapid identification of patients with primary EBV. infection and those with EBV infectious mononucleosis.
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页码:736 / 739
页数:4
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