Rapid identification of Beijing measles vaccine virus and wild virus by multiplex real-time fluorescent PCR

Abstract

To identify wild measles virus and vaccine virus by detection nucleic acid of clinical samples from measles patients with immunization history circulating in Beijing through multiplex real-time fluorescent PCR technology. From July 2011 to February 2012, 10 throat swabs and 15 urine specimens were collected from 16 suspected measles patients who were 8 - 9 months old infants with immunization history in Beijing. The specificity of multiplex real-time fluorescent PCR was firstly tested by measles vaccine virus, wild virus and other respiratory virus. Then the vaccine virus and wild virus were titrated and diluted to test the sensitivity of the PCR method. The result was then compared with it analyzed by PCR-RFLP method. Meanwhile, the clinical sample of the measles patients were tested and confirmed by sequencing method. The primer-probe sets of Fam, Joe and Cy5 showed great specificity of measles virus, and could distinguish the measles vaccine virus and wild virus well. The sensitivity of this method to detect measles vaccine virus reached 0.1 CCID(50)/0.1 ml; and the sensitivity to wild virus reached 0.006 CCID(50)/0.1 ml; which were both higher than the sensitivity of PCR-RFLP method. Out of the 16 measles patients with vaccination history, 3 were negative and the other 13 all belonged to measles virus genotype A, and were confirmed to be vaccine virus by sequencing method. Multiplex real-time PCR method is accurate, rapid and sensitive to identify measles vaccine virus and wild virus. The method could greatly help us to find measles patients and to identify the vaccine-related cases.