As the coverage rate of the measles vaccine increases, not all patients present the typical symptoms of measles after exposure to the measles virus (MV). The virus loads in clinical specimens from patients with vaccine-modified non-typical measles are expected to be low compared with those of primary MV infection. A rapid and sensitive laboratory procedure is required for diagnosis of measles.
SYBR Green (TaKaRa) and TaqMan (ABI) real-time reverse transcription-polymerase chain reaction (RT-PCR) assays were developed to detect MV-RNA. For the real-time RT-PCR, primer sets were designed from a region of the MV H gene of the Edmonston strain (genotype A). A TaqMan probe specific for the H gene of genotype D MV was used. The minimum detectable level of MV-RNA in the SYBR Green and TaqMan real-time RT-PCR assays was evaluated using synthetic MV-RNA. The sensitivity of real-time RT-PCR was compared with that of nested RT-PCR and the virus isolation method using throat swabs and peripheral blood samples from patients with measles.
The minimum detectable level of RNA was 10 and 10(2) copies for SYBR Green RT-PCR and TaqMan RT-PCR, respectively. Ten-10(6) copies of standard RNA were linearly detected on SYBR Green RT-PCR. The sensitivity of SYBR Green RT-PCR was equal to that of nested RT-PCR. MV-RNA was detected in virus isolation-negative throat swabs on SYBR Green RT-PCR.
SYBR Green RT-PCR is a highly sensitive, rapid, and useful diagnostic procedure for the detection of MV.
"MV-RNA can be detected in a large variety of samples, including blood, throat swabs, nasopharyngeal aspirates, urine, and oral fluid, an alternative non-invasive, easy to collect sample, recommended by the WHO for the European measles elimination program [CDC, 2008; Hutse et al., 2010; Rota et al., 2011]. Although numerous molecular techniques for MV- RNA detection have been described [van Binnendijk et al., 2003; Ozoemena et al., 2004; Tischer et al., 2004; El Mubarak et al., 2005; Fujino et al., 2005; Hummel et al., 2006; Hubschen et al., 2008; Akiyoshi et al., 2010; Ito et al., 2010; Rota et al., 2011], few studies have evaluated the combined use of serological and molecular methods for measles diagnosis [van Binnendijk et al., 2003; Tischer et al., 2004; Hyde et al., 2009; Sanz et al., 2010]. Only a small number of studies have evaluated the use of molecular techniques in the context of MV outbreaks [Mosquera et al., 2005] and data concerning MV-RNA detection in oral fluid samples are limited [van Binnendijk et al., 2003; Hutse et al., 2010]. "
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