基于聚合酶链反应的限制性片段长度多态性（RFLP）与多重聚合酶链反应在人巨细胞病毒糖蛋白B（gB）基因分型中的比较效能

Comparative efficacy of PCR-based restriction fragment length polymorphism (RFLP) & multiplex PCR for glycoprotein B (gB) genotyping of human cytomegalovirus.

作者信息

Sowmya P, Dhanya V, Madhavan H N, Therese K L

机构信息

L & T Microbiology Research Centre, Vision Research Foundation, Chennai, India.

出版信息

Indian J Med Res. 2007 Aug;126(2):122-7.

PMID:17932436

Abstract

BACKGROUND & OBJECTIVE: Glycoprotein B (gB), involved in cell-to-cell transmission of human cytomegalovirus (HCMV), is a critical factor in tissue tropism and viral pathogenesis. The aim of the present study was to compare the efficiency of PCR-based RFLP and multiplex nested PCR for gB gene of HCMV to determine their genotype in clinical specimens from patients with HCMV.

METHODS

The PCR based RFLP and the multiplex nested PCR were applied on standard strain of HCMV AD169, 4 clinical HCMV isolates and 70 clinical specimens positive for HCMV by pp65 antigenaemia assay or nested PCR for mtr II region or both.

RESULTS

Three of the four clinical isolates were genotyped as gB1 and the other as gB3 by both the methods. HCMV genome in all the 70 clinical specimens were genotyped by multiplex nested PCR whereas only 65 were genotyped by PCR-based RFLP. Forty one of 65 clinical specimens, gave concordant results by both methods. In the remaining 24, mixed infection with multiple genotypes was identified by multiplex nested PCR whereas single genotypes were identified by PCR-based RFLP.

INTERPRETATION & CONCLUSION: Multiplex nested PCR provided a rapid, sensitive and cost-effective assay for gB genotyping of HCMV and allowed detection of multiple gB genotypes of HCMV in clinical samples compared to PCR-based RFLP.

摘要

背景与目的

糖蛋白B（gB）参与人巨细胞病毒（HCMV）的细胞间传播，是组织嗜性和病毒发病机制中的关键因素。本研究旨在比较基于PCR的限制性片段长度多态性分析（RFLP）和多重巢式PCR对HCMV gB基因的检测效率，以确定HCMV感染患者临床标本中的基因型。

方法

将基于PCR的RFLP和多重巢式PCR应用于HCMV AD169标准株、4株临床HCMV分离株以及70份通过pp65抗原血症检测或mtr II区域巢式PCR或两者均阳性的HCMV临床标本。

结果

两种方法均将4株临床分离株中的3株基因型鉴定为gB1，另一株为gB3。所有70份临床标本中的HCMV基因组均通过多重巢式PCR进行了基因分型，而基于PCR的RFLP仅对65份进行了基因分型。65份临床标本中的41份，两种方法结果一致。在其余24份标本中，多重巢式PCR鉴定为多种基因型的混合感染，而基于PCR的RFLP鉴定为单一基因型。