Droplet Digital PCR (ddPCR) Does Not Enhance the Sensitivity of Detection of Cytomegalovirus (CMV) DNA in Newborn Dried Blood Spots Evaluated in the Context of Newborn Congenital CMV (cCMV) Screening

IF 4 Q1 GENETICS & HEREDITY
Nelmary Hernandez-Alvarado, Craig J. Bierle, M. Schleiss
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Abstract

Congenital cytomegalovirus (cCMV) infection is a leading cause of sensorineural hearing loss (SNHL) and neurodevelopmental disabilities in children worldwide. Some regions in the United States and Canada have implemented universal newborn screening for cCMV, which requires molecular diagnostic technologies for identifying cCMV, such as PCR testing of newborn dried blood spots (DBS). This study aimed to evaluate the sensitivity of droplet digital PCR (ddPCR) compared to quantitative real-time PCR to detect CMV DNA in newborn DBS. The limit of detection of various ddPCR primer/probe combinations (singleplex UL55-HEX, singleplex UL83-FAM, and multiplex UL55-HEX/UL83-FAM) was evaluated using the National Institute of Standards and Technology’s (NIST) CMV quantitative standard. Singleplex UL55-HEX ddPCR exhibited the lowest limit of detection among the primer/probe combinations tested for ddPCR. UL55 ddPCR was then compared to real-time PCR in 49 infants with confirmed cCMV identified through newborn screening for CMV in saliva swabs and confirmed by a urine test. The results showed that ddPCR was only positive for 59% (29 out of 49) of the cCMV infants, while real-time PCR was positive for 80% (39 out of 49). Due to its lower sensitivity and throughput, ddPCR may not be suitable for cCMV newborn screening.
在新生儿先天性巨细胞病毒 (cCMV) 筛查中评估液滴数字 PCR (ddPCR) 是否能提高新生儿干血斑中巨细胞病毒 (CMV) DNA 的检测灵敏度
先天性巨细胞病毒(cCMV)感染是导致全球儿童感音神经性听力损失(SNHL)和神经发育障碍的主要原因。美国和加拿大的一些地区已开始普及新生儿巨细胞病毒筛查,这需要采用分子诊断技术来识别巨细胞病毒,如新生儿干血斑(DBS)的 PCR 检测。本研究旨在评估与定量实时 PCR 相比,液滴数字 PCR(ddPCR)检测新生儿干血斑中 CMV DNA 的灵敏度。使用美国国家标准与技术研究院(NIST)的 CMV 定量标准评估了各种 ddPCR 引物/探针组合(单倍 UL55-HEX、单倍 UL83-FAM 和多倍 UL55-HEX/UL83-FAM)的检测限。在进行 ddPCR 测试的引物/探针组合中,单重 UL55-HEX ddPCR 的检测限最低。随后,UL55 ddPCR 与实时 PCR 对 49 名通过新生儿唾液拭子中的 CMV 筛查发现并通过尿液检测确诊为 cCMV 的婴儿进行了比较。结果显示,ddPCR 仅对 59% 的 cCMV 婴儿(49 人中有 29 人)呈阳性,而实时 PCR 则对 80% 的 cCMV 婴儿(49 人中有 39 人)呈阳性。由于 ddPCR 的灵敏度和通量较低,因此可能不适合用于 cCMV 新生儿筛查。
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来源期刊
International Journal of Neonatal Screening
International Journal of Neonatal Screening Medicine-Pediatrics, Perinatology and Child Health
CiteScore
6.70
自引率
20.00%
发文量
56
审稿时长
11 weeks
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