인간거대세포바이러스 DNA 정량 중합효소연쇄반응 검사의 실제
Practical Aspects of Cytomegalovirus DNA Quantitative PCR
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인간거대세포바이러스(human cytomegalovirus, CMV)는 면역이 저하된 환자에서 심각한 감염을 일으킬 수 있다. 장기 이식 수혜자에서 CMV 감염의 선제치료를 위해 일반적으로 CMV DNA 정량검사를 실시한다. 2006년 WHO에서 핵산증폭 검사를 위한 CMV 표준물질을 만들었지만, CMV DNA 정량 결과에는 검체, 핵산 추출법, 시발체와 소식자 및 증폭 반응의 효율성, 그리고 기기의 종류 등이 영향을 줄 수 있음을 고려해야 한다. CMV 정량검사의 검체는 주로 전혈 또는 혈장, 그리고 혈청이나 백혈구 분획을 이용하는데, 검체를 선택할 때 전혈과 백혈구 분획이 혈장이나 혈청보다 CMV DNA 검출 민감도가 높다는 점을 고려해야 한다. 임상 검사실에서 주로 사용되는 핵산 추출 방법은 대부분 흡착 크로마토그래피법의 원리를 이용한다. 그러나 핵산 추출 효율은 수기법과 자동화법에서 다를 수 있으며 추출 키트마다 차이가 있을 수 있다. 가장 널리 사용되는 CMV DNA 정량 PCR 검사는 실시간 중합효소연쇄반응을 이용한 것으로 이는 직선성 구간이 넓고, 검출 한계와 정량 한계값이 낮으며, 증폭 산물에 의한 오염 위험성이 낮다. 다른 진단검사와 마찬가지로 CMV DNA 정량 PCR 검사도 분석 전, 분석, 분석 후 전 과정에 대한 정도관리가 필요하다. 추적 관찰 간격은 5-7일이 적당하며 적절한 치료에도 반응이 없는 경우 약제 내성 CMV의 출현을 의심해야 한다. 선제치료를 위한 CMV 정량검사에서 하나의 검체를 선택하여 추적 관찰하며, WHO 표준물질에 맞춘 정량 PCR법을 사용하도록 한다. 향후 표준화된 검사법과 임상적으로 의미 있는 CMV DNA 농도에 대한 연구가 필요하다. [Ann Clin Microbiol 2017;20:21-26]
Abstract
Human cytomegalovirus (CMV) is a clinically important pathogen that causes significant morbidity and mortality in immunocompromised patients and is typically monitored using real-time polymerase chain reaction (real-time PCR). International standards and certified reference materials were recently developed by the WHO, providing the opportunity to standardize viral load reporting. Clinical microbiologists who conduct quantitative CMV DNA testing should be aware of technical issues that can affect the analytical and clinical performance of the method used. These include specimen type, limits of detection and quantification, linear range, reproducibility, and wide variability in viral load values among different assays. Specimen types for testing include whole blood, plasma, serum, and peripheral blood leukocytes. The tests that use whole blood and peripheral blood leukocytes have higher sensitivities. Adsorption chromatography methods are widely used for nucleic acid extraction, and efficiencies can differ between manual and automatic processes. The most common method for quantitative CMV DNA testing is real-time PCR. All CMV testing methods require quality control at the pre-analytic, analytic, and post-analytic stages. In transplant patients, specific quantitative results and monitoring of any changes at follow-up are important. Five to seven days is an adequate follow-up interval in this regard, and drug-resistant CMV should be suspected if there is no response to therapy. One specimen type should be chosen for follow-up quantitative CMV DNA testing, optimized according to WHO standards. Further studies are needed to better standardize CMV testing approaches and to determine the appropriate clinical cut-off level. (Ann Clin Microbiol 2017;20:-26)
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