We made use of specimens from a large, well-characterized mumps outbreak to assess the sensitivity of diagnostic tests for mumps. Thus, we were able to evaluate the impact of vaccination status and the timing of specimen collection on the relative abilities of the standard diagnostic methods (IgM versus RNA detection) to confirm mumps infection as well as the performance of different IgM assays. The methodology for mumps rRT-PCR was improved by both the availability of a large number of clinical specimens from confirmed cases and the quality of the specimens, as demonstrated by the high proportion that were positive by virus isolation.
All serologic assays in the evaluation performed well to confirm the few cases in unvaccinated individuals, but the two capture IgM assays detected IgM in a higher percentage of the cases with a history of vaccination than the indirect EIA and IFA. Although an improvement in sensitivity was noted for all assays when serum was collected ≥3 days after onset, the effect of timing of collection did not alter the overall higher detection by the capture IgM assays. While the Microimmune capture IgM assay performed the best among the commercial assays, a higher percentage of IgM-positive samples was obtained with the CDC capture IgM assay. The apparent higher rate of detection of IgM by the CDC assay than the commercial capture IgM test could be due to selection of the serum specimens chosen to evaluate and validate the tests. Although avidity testing has revealed that persons with a secondary immune response are capable of eliciting an IgM response, the IgM response appears to be variable and generally not as reactive in EIAs as that observed in a typical primary response (20
). Serum panels used to set the parameters for IgM detection (sensitivity) during the development of commercial assays are more likely to be collected from persons having a primary immune response and therefore have higher levels of IgM.
The timing of serum collection and the selection of the assay are both important when the majority of cases have been previously vaccinated. During a mumps outbreak in Nova Scotia, collection of convalescent phase serum from 25 mumps cases yielded only one additional confirmation by IgM using an indirect EIA (23
). While use of a capture IgM EIA may have improved IgM detection in that situation, individuals who have previously responded to mumps vaccination may not produce mumps-specific IgM, or the levels present may be undetectable by any assay.
The outbreak in New York City also provided the opportunity to discern differences in sensitivity in rRT-PCR using N versus SH as the target gene. Among 296 cases, rRT-PCR-N increased RNA detection by 14% compared with that of rRT-PCR-SH. There were positive results for 209 (71%) samples using the rRT-PCR-N. An equal number of samples were positive by virus isolation. However, 12 samples that were negative by virus isolation were positive by rRT-PCR-N, and 12 that were negative by rRT-PCR-N were positive by virus isolation. Buccal swab samples from 69 additional cases of mumps in New York City from January to March 2010 continued to be tested by both rRT-PCR-N and virus isolation. Concordant results were obtained by rRT-PCR-N and virus isolation with the exception of 4 samples that were again equally divided for positive results. Therefore, in most cases, rRT-PCR should provide sensitivity equivalent to viral culture and with much more rapid results.
A higher rate of case confirmation was obtained in the New York City outbreak by both IgM detection and by rRT-PCR than had been described following mumps outbreaks in 2006 using the same methods at the CDC (9
). Mumps RNA was detected in 57% of the cases in New York City using rRT-PCR-SH, an increase of over 25% compared to that obtained from 2 outbreaks in 2006 (9
). The proportion of cases that were positive by the CDC capture IgM EIA was also higher in this study (50%) than that of the outbreaks in 2006 (13% to 14%), even though the proportions of vaccinated cases were comparable (7
). Consistent with previous reports, the sensitivity obtained by rRT-PCR was higher than that obtained for IgM detection. However, this study demonstrated that the timing of sample collection and the type of IgM assay utilized are important factors.
There are several possible explanations for the improved rates of case confirmation in the New York City outbreak compared to those in the previous reports in which the same laboratory testing methods were used. The number of non-mumps cases or cases with mild symptoms included in the evaluation of the outbreaks in 2006 may have been much higher. Whereas the cases in New York City were nearly all described as having parotitis, it was not unusual for symptoms to be recorded as jaw or facial soreness or swelling during the outbreaks in 2006 (8
) (CDC, unpublished data). The intensity of exposure among the boys who attended Jewish schools was suggested to have played a large role in the New York City outbreak (13
). Lengthy study sessions involving face-to-face recitations provided opportunities for repeated and efficient transmission of infectious droplets. The clinical presentation of mumps among previously vaccinated persons may vary depending on the dose of virus transmitted. One could speculate that confirmation of mumps by RNA or IgM detection is more challenging among milder cases of mumps. Finally, adherence to recommendations for sample collection and transportation during the New York City outbreak may have contributed to the higher rate of laboratory confirmation observed in this study.
Our study has limitations. The small number of unvaccinated cases in the study and the unknown proportion of primary vaccine failures among the vaccinated cases may have limited our ability to ascertain the extent of variation in the performance of the assays based on vaccination status. Though the vaccinated cases were presumed to be the result of secondary vaccine failure, IgG avidity testing was not conducted. However, whereas all of the IgM assays detected IgM in 80% to 90% of the unvaccinated cases, IgM detection dropped to 10% to 51% among the vaccinated cases. While there may have been primary vaccine failures among the vaccinated cases, the lower sensitivity for IgM suggests that a majority of the vaccinated cases had a secondary type immune response. However, if the proportion of primary vaccine failures was higher among the New York City cases than among the vaccinated cases tested in the 2006 outbreaks, the difference could explain why we obtained higher rates of IgM and RNA positivity in this study.
The data from our study indicated that RNA detection by rRT-PCR decreases after 2 days after onset regardless of vaccination status. The low proportion of unvaccinated cases in this study may have limited our ability to detect an effect of vaccination status on the performance of rRT-PCR. The importance of early collection of samples for successful detection of mumps RNA in highly vaccinated populations has been reported previously and was speculated to be due to more rapid clearance of virus by persons with residual antibody from vaccination (9
The confirmed cases of mumps that were negative by all laboratory tests (n
= 42) may have been misclassified despite epidemiologic links to the outbreak. Inclusion of non-mumps cases would result in lower sensitivity of the diagnostic assays. However, the localized nature of the New York City outbreak (13
) and the improvement in PCR and/or IgM detection compared to the results obtained in 2006 suggest that most, if not all, of the cases were true cases of mumps.
It is recommended that providers collect both buccal swab specimens and serum samples when mumps infection is suspected. Although it was demonstrated that rRT-PCR was more sensitive than IgM detection, particularly when samples were collected within 2 days of symptom onset, the quality of virologic samples is variable, and delays in obtaining specimens are often unavoidable. Provider education regarding the proper collection techniques, transport methods, and recommended assays may improve the ability to confirm mumps infection among vaccinated populations.