Co-detection of the measles vaccine and wild-type virus by real-time PCR: public health laboratory protocol.

In rare cases vaccination with the measles virus vaccine genotype A (MeVA) may cause a vaccine reaction with clinical signs similar to infection with wild-type measles virus (MeVwt). Rapid differentiation between MeVA and MeVwt infection is important for taking adequate public health measures. Recently, a few MeVA real-time reverse-transcription quantitative PCR methods (RT-qPCRs) were described that can distinguish between MeVA and MeVwt. However, detection of MeVA does in theory not exclude infection with MeVwt. In the present study, we established a protocol for determination of co-infections with MeVA and MeVwt. To this end, MeVA RT-qPCRs were used in combination with the routine measles virus (MeV) RT-qPCR, and the results suggested that the differences between the RT-qPCR Ct values (delta Ct, ∆Ct) could be used as criteria. Subsequently, we tested samples from vaccine-associated measles cases that were confirmed by genotyping. In addition, experimental mixtures of MeVA and MeVwt were tested in different concentrations. All tested MeVA clinical samples had ∆Ct ≤3.6. The results of experimental mixtures showed a mean ∆Ct ≤2.8 for genotype A alone and >3.2 when combined with either genotype B3 or D8. The results of a receiver operator characteristic analysis indicated that the optimum ∆Ct for use as a cut-off value was 3.5, while with ∆Ct values of 2.9 and 3.7 sensitivity and specificity were respectively 1.00. Thus, ∆Ct could be used to exclude the presence of MeVwt if MeVA is detected and ∆Ct is <2.9, while ∆Ct >3.7 were highly suggestive of co-infection and ≥2.9 ∆Ct <3.7 warranted additional confirmation, such as next-generation sequencing. This RT-qPCR-based protocol could be used for the exclusion of infection with MeVwt in cases with vaccine-associated measles reaction, crucial for the timely implementation of public health prevention and control measures.

Emergent measles-containing vaccination recommendation for aged 6-11 months and detection of vaccine-associated measles during a large measles outbreak in Okinawa, Japan, in 2018.

Okinawa Prefecture, Japan, experienced a large measles outbreak from March to May 2018. During this outbreak, there were 99 laboratory-confirmed cases and 14 vaccine-associated measles cases. In addition to the reinforcement of routine immunization, Okinawa prefectural government introduced emergent measles-containing vaccination recommendations for infants aged 6-11 months as part of the outbreak response. Increased concern exists in Okinawa about measles in infants following a previous outbreak from 1998 to 2001, when nine children including four infants died. Of 8062 infants aged 6-11 months who received measles-containing vaccine (MCV), six developed vaccine-associated measles; incidence was 0.74 per 1000 doses (95%CI 0.27-1.62). This was similar to that of first dose routine immunization recipients at one year of age (IR 0.60, 95%CI 0.20-1.78). Among 14 vaccine-associated measles cases, throat swab samples showed the highest positive rate (92.9%) by real-time reverse transcription polymerase chain reaction (RT-qPCR), followed by urine (25.0%) and whole blood (7.7%) samples. Furthermore, one throat swab sample classified as equivocal by RT-qPCR was positive by conventional RT-PCR (RT-PCR). During an outbreak, it is critical to distinguish between cases with measles-like symptoms caused by wild circulating virus and those caused by vaccine-derived virus as accurately and urgently as possible because the public health response will be quite different. No infant deaths were observed during this outbreak, and no severe adverse events following immunization were seen among infants 6-11 months old who were given MCV as a public health response. Thus, we conclude that introduction of emergent MCV was effective and describing the characteristics of vaccine-associated measles cases during a measles outbreak will be helpful for future outbreak response efforts.

Laboratory diagnosis of vaccine-associated measles in Zhejiang Province, China.

BACKGROUND/PURPOSE: Along with the improving vaccine coverage, suspected vaccine-associated measles has been reported in Zhejiang Province, China. In order to maintain the accuracy of the measles surveillance system, it is critical to discriminate between measles vaccine and wild-type virus. METHODS: Eight suspected cases of vaccine-associated measles were reported in Zhejiang Province during 2011 and 2014. Sera collected within 4 days and throat swabs collected within 6 days after rash onset were tested with immunoglobulin M and measles virus (MeV) RNA to confirm MeV infection. In order to further identify the vaccine-associated cases, throat swabs with positive MeV RNA were tested using an allelic discrimination real-time reverse transcriptase polymerase chain reaction (rRT-PCR) assay developed in this study, RT-PCR-restriction fragment length polymorphism (RFLP) recommended by the National Measles Laboratory, and RT-PCR followed by sequencing and genotyping. RESULTS: Combining anti-measles immunoglobulin M and RNA testing, eight cases were confirmed as MeV infection. Of the eight, two were identified as vaccine-associated cases by the allelic discrimination rRT-PCR assay, and one was identified by RT-PCR-RFLP. Subsequent sequencing and genotyping confirmed that the sequences of the two cases were identical to that of the Chinese vaccine strain. The developed allelic discrimination rRT-PCR was 10 times more sensitive than the RT-PCR-RFLP assay when RNA standards generated from three genotypes of MeV were tested. CONCLUSION: Vaccine-associated measles has been identified in Zhejiang. The developed allelic discrimination rRT-PCR assay is rapid and sensitive, which will facilitate the surveillance for vaccine-associated measles.