Ann Lab Med.  2017 Mar;37(2):129-136. 10.3343/alm.2017.37.2.129.

Automated Nucleic Acid Extraction Systems for Detecting Cytomegalovirus and Epstein-Barr Virus Using Real-Time PCR: A Comparison Study Between the QIAsymphony RGQ and QIAcube Systems

• Kim H
• Hur M
• Kim JY
• Moon HW
• Yun YM
• Cho HC

• KMID: 2373630
• DOI: http://doi.org/10.3343/alm.2017.37.2.129

Abstract

BACKGROUND
Cytomegalovirus (CMV) and Epstein-Barr virus (EBV) are increasingly important in immunocompromised patients. Nucleic acid extraction methods could affect the results of viral nucleic acid amplification tests. We compared two automated nucleic acid extraction systems for detecting CMV and EBV using real-time PCR assays.
METHODS
One hundred and fifty-three whole blood (WB) samples were tested for CMV detection, and 117 WB samples were tested for EBV detection. Viral nucleic acid was extracted in parallel by using QIAsymphony RGQ and QIAcube (Qiagen GmbH, Germany), and real-time PCR assays for CMV and EBV were performed with a Rotor-Gene Q real-time PCR cycler (Qiagen). Detection rates for CMV and EBV were compared, and agreements between the two systems were analyzed.
RESULTS
The detection rate of CMV and EBV differed significantly between the QIAsymphony RGQ and QIAcube systems (CMV, 59.5% [91/153] vs 43.8% [67/153], P=0.0005; EBV, 59.0% [69/117] vs 42.7% [50/117], P=0.0008). The two systems showed moderate agreement for CMV and EBV detection (kappa=0.43 and 0.52, respectively). QIAsymphony RGQ showed a negligible correlation with QIAcube for quantitative EBV detection. QIAcube exhibited EBV PCR inhibition in 23.9% (28/117) of samples.
CONCLUSIONS
Automated nucleic acid extraction systems have different performances and significantly affect the detection of viral pathogens. The QIAsymphony RGQ system appears to be superior to the QIAcube system for detecting CMV and EBV. A suitable sample preparation system should be considered for optimized nucleic acid amplification in clinical laboratories.

Keyword

Cytomegalovirus; Epstein-Barr virus; QIAsymphony RGQ; QIAcube; Nucleic acid; Extraction; Performance

MeSH Terms

Automation
Cytomegalovirus/*genetics/isolation & purification
Cytomegalovirus Infections/diagnosis/*virology
DNA, Viral/*blood/isolation & purification/metabolism
Herpesvirus 4, Human/*genetics/isolation & purification
Humans
Reagent Kits, Diagnostic
Real-Time Polymerase Chain Reaction
DNA, Viral
Reagent Kits, Diagnostic

Figure

• 

  Fig. 1 Linearity of quantification using 1:10 serial dilutions of the first WHO international standards for (A) cytomegalovirus (CMV) and (B) Epstein-Barr virus (EBV). Viral load results were obtained using the artus CMV QS-RGQ Kit and the artus EBV QS-RGQ kit in combination with the Rotor Gene Q thermal cycler. The “0” results (N=2 for CMV and N=1 for EBV) are not demonstrated in this figure because of the log transformation. Solid lines indicate the linear regression fit.

• 

  Fig. 2 Discrepant cytomegalovirus (CMV, N=44; left panel) and Epstein-Barr virus (EBV, N=29; right panel) results using the QIAsymphony RGQ and QIAcube systems.

• 

  Fig. 3 Comparison of cytomegalovirus (CMV, N=38) and Epstein-Barr virus (EBV, N=17) results using the QIAsymphony RGQ and QIA-cube systems. (A) Passing-Bablok regression for CMV detection. (B) Bland-Altman plot for CMV detection. (C) Passing-Bablok regression for EBV detection. (D) Bland-Altman plot for EBV detection. In the regression plot, the solid line indicates the regression line and dashed lines indicate 95% confidence interval. In the Bland-Altman plot, the bold line indicates the mean difference between values, the dashed lines indicate 95% confidence interval, and the solid lines indicate mean difference ±1.96 standard deviation.

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