In the past decade, multiple mumps outbreaks have occurred in the United States, primarily in close-contact, high-density settings such as colleges, with a high attack rate among young adults, many of whom had the recommended 2 doses of mumps-measles-rubella (MMR) vaccine. Waning humoral immunity and the circulation of divergent wild-type mumps strains have been proposed as contributing factors to mumps resurgence. Blood samples from 71 healthy 18- to 23-year-old college students living in a non-outbreak area were assayed for antibodies and memory B cells (MBCs) to mumps, measles, and rubella. Seroprevalence rates of mumps, measles, and rubella determined by IgG enzyme-linked immunosorbent assay (ELISA) were 93, 93, and 100%, respectively. The index standard ratio indicated that the concentration of IgG was significantly lower for mumps than rubella. High IgG avidity to mumps Enders strain was detected in sera of 59/71 participants who had sufficient IgG levels. The frequency of circulating mumps-specific MBCs was 5 to 10 times lower than measles and rubella, and 10% of the participants had no detectable MBCs to mumps. Geometric mean neutralizing antibody titers (GMTs) by plaque reduction neutralization to the predominant circulating wild-type mumps strain (genotype G) were 6-fold lower than the GMTs against the Jeryl Lynn vaccine strain (genotype A). The majority of the participants (80%) received their second MMR vaccine >/=10 years prior to study participation. Additional efforts are needed to fully characterize B and T cell immune responses to mumps vaccine and to develop strategies to improve the quality and durability of vaccine-induced immunity.

Introduction: Mumps is a vaccine-preventable viral disease that may cause deafness, orchitis, encephalitis or death. However, mumps vaccine is not included in Zambia's Expanded Program for Immunization. In January 2015, Integrated Disease Surveillance and Response data revealed an increase in reported mumps cases in Luanshya District. We investigated to confirm the etiology and generate epidemiological data on mumps in Zambia. Method(s): We conducted active case finding, examined possible case-patients, and administered a standard questionnaire. A suspected mumps case was defined as acute onset of salivary gland swelling in a Luanshya resident during January-June 2015. Eight case-patients provided serum samples to test for mumps-specific immunoglobulin IgM, and buccal swabs to test for mumps viral RNA by RT-PCR, and genotyping of mumps virus at the Centers for Disease Control and Prevention, Atlanta, Georgia, USA. Result(s): From January - June 2015, a total of 283 mumps cases were reported in Luanshya, peaking in April (71 cases) and clustering (81%) in two townships. Of 72 suspected case-patients interviewed, 81% were aged < 15 years (29%, 1-4 years) and 61% were female. Common clinical characteristics were buccal tenderness (29%) and fever > 37.5C (29%). Mumps virus genotype D was confirmed in five case-patients who tested positive by RT-PCR; six case-patients were sero-positive for anti-mumps IgM antibodies (total seven lab-confirmed cases). Conclusion(s): Our findings represent the first reported epidemiologic description of mumps in Zambia. While the epidemiology is consistent with prior descriptions of mumps in unimmunized populations and no serious complications arose, this report provides data to inform policy discussions regarding mumps vaccination in Zambia.

Waning mumps IgG antibody and incomplete IgG avidity maturation may increase susceptibility to mumps virus infection in some vaccinees. To measure mumps IgG avidity, serum specimens serially diluted to the endpoint were incubated on a commercial mumps-specific IgG enzyme immunoassay and treated with the protein denaturant diethylamine (60 mM, pH 10). End titer avidity indices (etAIs [percent ratio of detected diethylamine-resistant IgG at endpoint]) were calculated. Unpaired serum specimens (n = 108) from 15-month-old children living in a low-incidence setting were collected 1 month and 2 years after the first measles, mumps, and rubella vaccine dose (MMR1) and tested for mumps avidity. Per the receiver operating characteristic curve, the avidity assay is accurate (area under the curve, 0.994; 95% confidence interval [CI], 0.956 to 1.000), 96.5% sensitive (95% CI, 87.9 to 99.6%), and 92.2% specific (95% CI, 81.1 to 97.8%) at an etAI of 30%. When 9 sets of paired serum specimens collected 1 to 60 months post-MMR1 were tested for mumps and measles IgG avidity using comparable methods, the mumps etAI increased from 11% to 40 to 60% in 6 months. From 6 to 60 months, avidity was sustained at a mean etAI of 50% (95% CI, 46 to 54%), significantly lower (P < 0.0001) than the mean measles etAI of 80% (95% CI, 74 to 86%). Mean etAIs in children 2 years post-MMR1 (n = 51), unvaccinated adults with distant mumps disease (n = 29), and confirmed mumps cases (n = 23) were 54, 62, and 57%, respectively. A mumps-specific endpoint avidity assay was developed and validated, and mumps avidity was determined to be generally sustained at etAIs of 40 to 60%, reaching etAIs of >80% in some individuals.IMPORTANCE Numerous outbreaks of mumps have occurred in the United States among two-dose measles-mumps-rubella (MMR)-vaccinated populations since 2006. The avidity of mumps-specific IgG antibodies may affect susceptibility to mumps virus infection in some vaccinated individuals. To accurately measure mumps avidity, we developed and validated a mumps-specific IgG avidity assay that determines avidity at the endpoint titer of serially diluted serum specimens, providing results that are independent of IgG concentration. At low antibody titers, endpoint methods are considered more accurate than methods that determine avidity at a single dilution. We determined that 6 months after the first MMR dose, mumps IgG avidity is high and generally sustained at avidity indices of 40 to 60%, reaching values of >80% in some individuals. Additionally, 4% (4/103) of individuals had avidity indices of </=30% (low avidity) 2 years after vaccination. Inadequate mumps avidity maturation may be one factor influencing susceptibility to mumps virus infection among previously vaccinated or naturally infected individuals.

Background: Recent mumps outbreaks among 2-dose measles mumps rubella (MMR) vaccine recipients have raised questions regarding the potential benefits of a third dose of vaccine (MMR3). If MMR3 provides a sustained elevation in mumps antibody, it may be beneficial for certain at-risk groups or as an outbreak control measure. Methods: Sera were collected immediately prior to MMR3 and at 1 month and 1 year post-MMR3 from 656 healthy adults aged 18-28 years in a nonoutbreak setting. Immunoglobulin G (IgG) was measured by enzyme-linked immunosorbent assay (ELISA) using whole mumps virus (commercial ELISA), hemagglutinin (HN; major neutralizing target), and nucleoprotein (NP; immunodominant) antigens. ELISA measurements were compared with in vitro plaque reduction neutralization (PRN) titers, and baseline antibody was compared with post-MMR3 levels. Results: There were modest but statistically significant (P < .05) increases in mumps antibody at 1 month post-MMR3 by all 3 ELISA methods and by PRN titer. At 1 year post-MMR3, mumps antibody declined toward baseline but remained elevated (P < .05). The correlation between PRN titers and ELISA measurements was poor (r(2) = .49), although sera with the highest amount of HN IgG also had the highest PRN titers. Conclusions: Individuals with the lowest baseline PRN titers had the largest increase in frequency of samples that became positive for HN and NP by ELISA. A third dose of MMR may benefit certain individuals with a low level of mumps virus-neutralizing antibody, especially in the context of an outbreak or other high-risk setting. Additionally, poor correlation among serologic tests does not allow effective prediction of PRN titer by ELISA.

In the United States, approximately 9% of the measles cases reported from 2012 to 2014 occurred in vaccinated individuals. Laboratory confirmation of measles in vaccinated individuals is challenging since IgM assays can give inconclusive results. Although a positive reverse transcription (RT)-PCR assay result from an appropriately timed specimen can provide confirmation, negative results may not rule out a highly suspicious case. Detection of high-avidity measles IgG in serum samples provides laboratory evidence of a past immunologic response to measles from natural infection or immunization. High concentrations of measles neutralizing antibody have been observed by plaque reduction neutralization (PRN) assays among confirmed measles cases with high-avidity IgG, referred to here as reinfection cases (RICs). In this study, we evaluated the utility of measuring levels of measles neutralizing antibody to distinguish RICs from noncases by receiver operating characteristic curve analysis. Single and paired serum samples with high-avidity measles IgG from suspected measles cases submitted to the CDC for routine surveillance were used for the analysis. The RICs were confirmed by a 4-fold rise in PRN titer or by RT-quantitative PCR (RT-qPCR) assay, while the noncases were negative by both assays. Discrimination accuracy was high with serum samples collected ≥3 days after rash onset (area under the curve, 0.953; 95% confidence interval [CI], 0.854 to 0.993). Measles neutralizing antibody concentrations of ≥40,000 mIU/ml identified RICs with 90% sensitivity (95% CI, 74 to 98%) and 100% specificity (95% CI, 82 to 100%). Therefore, when serological or RT-qPCR results are unavailable or inconclusive, suspected measles cases with high-avidity measles IgG can be confirmed as RICs by measles neutralizing antibody concentrations of ≥40,000 mIU/ml.

BACKGROUND: Two doses of measles-mumps-rubella (MMR) vaccine are 97% effective against measles, but waning antibody immunity and two-dose vaccine failures occur. We administered a third MMR dose (MMR3) to young adults and assessed immunogenicity over 1 year. METHODS: Measles virus (MeV) neutralizing antibody concentrations, cell-mediated immunity (CMI), and IgG antibody avidity were assessed at baseline, 1-month, and 1-year after MMR3. RESULTS: Of 662 subjects at baseline, 1 (0.2%) was seronegative (<8 mIU/mL) and 23 (3.5%) had low (8-120 mIU/mL) MeV neutralizing antibodies. At 1-month post-MMR3, 1 (0.2%) subject was seronegative and 6 (0.9%) had low neutralizing antibodies with only 21/662 (3.2%) showing a ≥4-fold rise in neutralizing antibodies. At 1-year post-MMR3, none were negative and 10 (1.6%) of 617 subjects had low neutralizing antibodies. CMI results showed low-levels of spot-forming cells after stimulation, suggesting T-cell memory, but the response was minimal post-MMR3. MeV IgG avidity results did not correlate with neutralization results. CONCLUSIONS: Most subjects were seropositive pre-MMR3 and very few had a secondary immune response post-MMR3. Similarly, CMI and avidity results showed minimal qualitative improvements in immune response post-MMR3. We did not find compelling data to support a routine third dose of MMR vaccine.

Neutralizing antibodies are assumed to be essential for protection against mumps virus infection, but their measurement is labor and time intensive. For this reason ELISA assays are typically used to measure mumps specific IgG. However, since there is poor correlation between mumps neutralization titers and ELISA assays that measure the presence of mumps specific IgG, ELISA assays that better correlate with neutralization are needed. To address this issue, we measured mumps antibody by plaque reduction neutralization, by a commercial ELISA (whole-virus antigen), and by ELISAs specific for the mumps nucleoprotein and hemagglutinin. The results indicate that differences in the antibody response to the individual mumps proteins could partially explain the lack of correlation among various serologic tests. Furthermore, the data indicate that some seropositive individuals have low levels of neutralizing antibody. If neutralizing antibody is important for protection, this suggests that previous estimates of immunity based on whole-virus ELISA may be overstated.

A mumps outbreak in upstate New York in 2009 at a summer camp for Orthodox Jewish boys spread into Orthodox Jewish communities in the Northeast including New York City. The availability of epidemiologic information including vaccination records and parotitis onset dates allowed an enhanced analysis of laboratory methods for mumps testing. Serum and buccal swab samples were collected from 296 confirmed cases with onsets from September through December, 2009. All samples were tested using the CDC capture IgM EIA and a real-time RT-PCR (rRT-PCR) that targets the short hydrophobic gene. A subset of the samples (n=205) was used to evaluate 3 commercial mumps IgM assays and to assess the sensitivity of using an alternative target gene (nucleoprotein) in the rRT-PCR protocol. Among 115 cases of mumps with 2 documented doses of measles, mumps, and rubella (MMR) vaccine, the CDC capture IgM EIA detected IgM in 51% of serum samples compared to 9%-24% using three commercial IgM assays. The rRT-PCR that targeted the nucleoprotein gene increased RNA detection by 14% compared to that obtained with the original protocol. The ability to detect IgM improved when serum was collected 3 days or more after onset whereas sensitivity of RNA detection by rRT-PCR declined when buccal swabs were collected later than 2 days after onset. Selection of testing methods and timing of sample collection are important factors in the ability to confirm infection among vaccinated persons. These results reinforce the need to use virus detection assays in addition to serologic tests.

Waning immunity or secondary vaccine failure (SVF) has been anticipated by some as a challenge to global measles elimination efforts. Although such cases are infrequent, measles virus (MeV) infection can occur in vaccinated individuals following intense and/or prolonged exposure to an infected individual and may present as a modified illness that is unrecognizable as measles outside of the context of a measles outbreak. The immunoglobulin M response in previously vaccinated individuals may be nominal or fleeting, and viral replication may be limited. As global elimination proceeds, additional methods for confirming modified measles cases may be needed to understand whether SVF cases contribute to continued measles virus (MeV) transmission. In this report, we describe clinical symptoms and laboratory results for unvaccinated individuals with acute measles and individuals with SVF identified during MeV outbreaks. SVF cases were characterized by the serological parameters of high-avidity antibodies and distinctively high levels of neutralizing antibody. These parameters may represent useful biomarkers for classification of SVF cases that previously could not be confirmed as such using routine laboratory diagnostic techniques.

Although high measles, mumps, and rubella (MMR) vaccination coverage has been successful in dramatically reducing mumps disease in the United States, mumps (re)infections occasionally occur in individuals who have been either previously vaccinated or naturally infected. Standard diagnostics that detect virus or virus-specific antibody are dependable for confirming primary mumps infection in immunologically naive persons, but these methods perform inconsistently on individuals with prior immune exposure. We hypothesized that detection of activated mumps-specific Antibody-Secreting B-Cells (ASC) by ELISpot could be used as a more reliable diagnostic. To test this, a time-course of virus-specific ASC responses was measured by ELISpot following MMR vaccination of 16 previously vaccinated or naturally exposed adult volunteers. Mumps-specific ASC were detectable in 68% of these individuals at some point during the first 3 weeks following re-vaccination. In addition, mumps-specific ASC were detected in 7/7 previously vaccinated individuals who recently had been infected as part of a confirmed mumps outbreak. These data suggest that ELISpot detection of mumps-specific ASC has the potential for use as an alternative method of diagnosis when suspect cases cannot be confirmed by detection of IgM or virus. In addition, it was determined that mumps-specific memory B-cells are detected at a much lower frequency than measles or rubella-specific cells, suggesting that mumps infection may not generate robust B-cell memory.