Combination of Reverse Transcriptase PCR Analysis and Immunoglobulin M Detection on Filter Paper Blood Samples Allows Diagnostic and Epidemiological Studies of Measles

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Abstract
As measles control and elimination campaigns progress, laboratory confirmation of clinically diagnosed measles cases becomes increasingly important. However, in many tropical countries collection and storage of clinical specimens for this purpose are logistically complicated. In this study it is shown that blood samples spotted on filter paper are suitable for the laboratory diagnosis of measles using a combination of reverse transcriptase PCR (RT-PCR) analysis and immunoglobulin M (IgM) detection. First, it was shown that in vitro measles virus (MV)-infected cells diluted in human blood and spotted on filter paper can be detected by RT-PCR. Small amounts of infected cells remained detectable after 25 weeks of storage of the filter paper at room temperature, 4 weeks at 37°C, or 2 weeks at 45°C. Subsequently, this RT-PCR was applied to filter paper blood samples collected from 117 clinically diagnosed measles patients in Sudan in 1997 and 1998. Prior laboratory diagnosis had confirmed 90 cases as acute MV infections, while 27 proved to be nonmeasles rash disease cases. Positive RT-PCR signals were detected in filter paper blood samples of 43 of the 90 confirmed cases (48%) but in none of the 27 nonmeasles cases. In addition, MV-specific IgM levels measured in reconstituted filter paper samples correlated well with those measured in plasma samples. Measles diagnosis based on the combination of filter paper RT-PCR and IgM detection had a sensitivity and specificity of 99 and 96%, respectively. An advantage of this diagnostic approach is that sequencing of RT-PCR products allows phylogenetic analysis of the MV strain involved.