Comparison of the Cytomegalovirus (CMV) Enzyme-Linked Immunosorbent Spot and CMV QuantiFERON Cell-Mediated Immune Assays in CMV-Seropositive and -Seronegative Pregnant and Nonpregnant Women

Abstract

Human cytomegalovirus (CMV) infection is a major cause of congenital infection leading to birth defects and sensorineural anomalies, including deafness. Recently, cell-mediated immunity (CMI) in pregnant women has been shown to correlate with congenital CMV transmission. In this study, two interferon gamma release assays (IGRA), the CMV enzyme-linked immunosorbent spot (ELISPOT) and CMV QuantiFERON assays, detecting CMV-specific CMI were compared. These assays were performed for 80 CMV-infected (57 primarily and 23 nonprimarily) pregnant women and 115 controls, including 89 healthy CMV-seropositive pregnant women without active CMV infection, 15 CMV-seronegative pregnant women, and 11 seropositive or seronegative nonpregnant women. Statistical tests, including frequency distribution analysis, nonparametric Kruskal-Wallis equality-of-populations rank test, Wilcoxon rank sum test for equality on unmatched data, and lowess smoothing local regression, were employed to determine statistical differences between groups and correlation between the assays. The CMV ELISPOT and CMV QuantiFERON assay data were not normally distributed and did not display equal variance. The CMV ELISPOT but not CMV QuantiFERON assay displayed significant higher values for primarily CMV-infected women than for the healthy seropositive pregnant and nonpregnant groups (P = 0.0057 and 0.0379, respectively) and those with nonprimary infections (P = 0.0104). The lowess local regression model comparing the assays on an individual basis showed a value bandwidth of 0.8. Both assays were highly accurate in discriminating CMV-seronegative pregnant women. The CMV ELISPOT assay was more effective than CMV-QuantiFERON in differentiating primary from the nonprimary infections. A substantial degree of variability exists between CMV ELISPOT and CMV QuantiFERON assay results for CMV-seropositive pregnant women.

FIG 1

Box-and-whisker distribution of CMV ELISPOT (A) and CMV QuantiFERON (B) assay values under different conditions, including pregnancy (preg), nonpregnancy (npreg), and CMV seropositivity (sero+ or sero−). “Primary” and “nonprimary” refer to CMV infection. CMV ELISPOT assay values were limited to 1,000 spots/2 × 10⁵ PBMCs, while CMV QuantiFERON assay values were limited to 10 IU of IFN-γ/ml. CMV viremia occurred prior to CMV ELISPOT and CMV QuantiFERON assay determination.

FIG 2

Frequency distribution of CMV ELISPOT (A) and CMV QuantiFERON (B).

FIG 3

Comparison of CMV ELISPOT and QuantiFERON in 195 pregnant and nonpregnant CMV-seropositive and -seronegative women. CMV ELISPOT values were limited to 1,000 spots/2 × 10⁵ PBMCs while CMV QuantiFERON was limited to 10 IU of IFN-γ/ml. The dashed line indicates the lowess local regression-smoothing model. The bandwidth is 0.8.