Development of a generic real-time PCR assay for simultaneous detection of proviral DNA of simian Betaretrovirus serotypes 1, 2, 3, 4 and 5 and secondary uniplex assays for specific serotype identification

开发通用实时 PCR 检测方法,用于同时检测猿猴 Beta 逆转录病毒血清型 1、2、3、4 和 5 的原病毒 DNA,以及用于特定血清型鉴定的二次单重检测

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作者:Jessica A White, Patricia A Todd, Ann N Rosenthal, JoAnn L Yee, Richard Grant, Nicholas W Lerche

Abstract

Simian betaretroviruses (formerly Type D retroviruses; SRV) are a group of closely related retroviruses for which the natural host species are Asian monkeys of the genus Macaca. Five serotypes have been identified by classical neutralization assays and three additional untyped variants have been reported (SRV(Tsukuba), SRV-6, SRV-7). These viruses may be significant pathogens in macaque colonies, causing a broad spectrum of clinical disease secondary to viral-induced immune suppression. Undetected SRV infections in research macaques also represent a potential confounding variable in research protocols and a concern for human caretakers. Intensive testing efforts have been implemented to identify infected animals in established colonies. A real-time quantitative generic multiplex PCR assay was developed that is capable of simultaneous detection of proviral DNA of SRV serotypes 1, 2, 3, 4 and 5. This assay incorporates amplification of the oncostatin M (OSM) gene for confirmation of amplifiable DNA and allows quantitation of the number of proviral copies per cell analyzed in each multiplex reaction. Detection of multiple serotypes by PCR increases the efficiency and cost-effectiveness of SRV screening programs. A panel of SRV serotype-specific uniplex real-time PCR assays for discrimination among the five recognized serotypes is also described.

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