Reevaluation of laboratory methods for diagnosis of measles.

The purpose of this study is to reevaluate the sensitivities of different methods used in the diagnosis of measles including virus isolation, RT-PCR, and measurement of IgM. Sixty-three throat swabs, 84 peripheral blood mononuclear cell (PBMC) samples, and 85 plasma samples were collected from 85 cases of suspected measles. The sensitivity of virus isolation using throat swabs and PBMC in comparison with RT-PCR was 58.1 and 93.5%, respectively. We defined laboratory-confirmed cases as those in which at least one of the methods was positive. The percentage of positive results by the different methods was compared among 49 laboratory-confirmed cases. The percentage of positive results from PBMC by RT-PCR and virus isolation was 100 and 91.7%, respectively. The percentage of positive results from throat swabs by RT-PCR and virus isolation was 91.2 and 52.8%, respectively. The percentage of IgM positive (79.6%) was significantly lower than that of PBMC by RT-PCR. Ten of 27 plasma samples collected within 5 days of the onset of fever were IgM negative. In contrast, all of the 21 plasma samples collected 6 days after the onset of fever were IgM positive. In conclusion, the detection of measles virus RNA in PBMC by RT-PCR was the most effective method for diagnosis of measles.

Detection of measles virus RNA on SYBR green real-time reverse transcription-polymerase chain reaction.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: SYBR Green RT-PCR is a highly sensitive, rapid, and useful diagnostic procedure for the detection of MV.