The burden of disease from measles has been greatly reduced by vaccination, and many countries have adopted a goal of measles elimination. To eliminate measles the administration of more than one dose of vaccine is necessary. Mass campaigns, in which measles vaccine is administered irrespective of prior receipt of vaccine or history of measles disease, are a major component of current programs to eliminate measles or enhance measles control. 1 Measles vaccine is administered by injection; thus there are potential risks associated with inappropriate reuse or unsafe disposal of used needles and syringes. 2 Previous studies have shown that the aerosol route is more immunogenic than injected vaccines in seronegative and seropositive children older than 9 months of age. 3 This route was used to vaccinate nearly 4 million schoolchildren in Mexico with apparently high effectiveness and safety. 4 Study of the duration of the increase in antibody after revaccination is important, because antibody wanes rapidly after revaccination by injection. 5, 6 We have recently demonstrated that the antibody responses of South African schoolchildren at 1 month and 1 year after revaccination were significantly higher after Edmonston-Zagreb (EZ) vaccine by aerosol than after EZ or Schwarz (SW) vaccine by subcutaneous injection. 7 We present data on the persistence of antibody at 2 years after revaccination and discuss the implications for the use of aerosol vaccination in sustaining humoral immunity to measles. Methods. Methods have been presented in detail elsewhere. 7 Briefly a randomized controlled Phase II vaccination trial was conducted in 4327 schoolchildren ages 5 through 14 years in Durban in 1996. Informed consent was obtained from parents or guardians of all children. Ethical approval for human experimentation was obtained from the University of Natal, South Africa, as well as from the London School of Hygiene and Tropical Medicine, UK. Children were allocated by block randomization of school classes to receive one of the following: EZ vaccine by aerosol (EZae), 5000 plaque-forming units (pfu)/dose; Schwarz vaccine by aerosol (SWae), 14 000 pfu/dose; EZ vaccine subcutaneously (EZsc), 10 000 pfu/dose; or Schwarz vaccine subcutaneously (SWsc), 12 000 pfu/dose. Sera were collected prevaccination, 1 month, 1 year and 2 years after revaccination and assayed by the hemagglutination inhibition (HI) assay. 8 The starting dilution was 1:4, corresponding to ∼300 mIU/ml which is slightly higher than the level considered to correlate with protection against measles disease. 9, 10 Data on clinical reactions to vaccines are presented elsewhere 7; no serious adverse events were detected. We had found that the Schwarz vaccine was inactivated during nebulization; hence results of children in this group are not presented here (these children were offered revaccination by subcutaneous injection). We followed up children in the other vaccine groups whose prevaccination titers were <1:4 and a random sample of those with higher baseline titers. Results. Of 992 children followed up to 1 year, 851 (86%) were recontacted at 2 years after revaccination. At baseline most children had a history of prior vaccination (n = 698) and/or measles-like illness (n = 268). An IgM response at 1 month after vaccination was detected by radioimmunoassay 11 in only 49 of 956 children. Prevaccination antibody titers were similar in all groups, ∼40% of children having titers <1:4. Among children with prevaccination antibody titers ≤1:8, most children showed a significant (≥4-fold) increase in titer at 1 month after revaccination, but titers then fell quite rapidly to 1 year. For all groups the rate of loss of antibody was greatest in the first year after revaccination, with geometric mean titers declining substantially more slowly in the second year (Fig. 1).Fig. 1: Hemagglutination inhibition titer changes after revaccination.The initial humoral response to revaccination was of greatest magnitude in the EZ aerosol group, and the differences between groups were sustained over time. By 2 years after revaccination only 6% of children in the EZae group had reverted to a titer below our cutoff of 1:4. The EZsc group was twice as likely (13%, P < 0.01), and the SWsc group three times as likely (19%, P < 0.001), to have titers below the threshold at this time. Seronegativity in the EZsc group was significantly less than in the SWsc group at 2 years (P = 0.037). The percentage of children whose antibody titer was at least 4-fold higher than baseline (“seroconversion”) was highest in the EZae group at all time points. In the EZae group 55% (176 of 323) had seroconverted at 2 years compared with only 23% (64 of 281) for EZsc and 21% (51 of 247) for SWsc (P < 0.001 for both comparisons). At 2 years the EZae group had the highest percentage of children with titers >1:8 (59.8%) compared with EZsc (29.2%) and SWsc (25.9%) (P < 0.001 for both comparisons). African children continued to have a lower response than Indian children at 2 years after revaccination after controlling for baseline antibody titer. However, African children receiving aerosol had significantly better seroconversion rates than the African children receiving subcutaneous vaccine at 1 month (80%vs. 62%, P = 0.005), at 1 year (52%vs. 26%, P < 0.001) and at 2 years (38%vs. 15%, P < 0.001) after revaccination. Furthermore African children vaccinated by aerosol had as good as or significantly better seroconversion rates than Indian children vaccinated by injection (at 1 month: 80%vs. 74%, P = 0.33; at 1 year: 52%vs. 31%, P < 0.001; at 2 years: 38%vs. 24%, P = 0.01). At 1 year after revaccination we had found significant differences in serologic response by several criteria when comparing EZ with SW by injection. At 2 years follow-up, however, the differences in seroresponse were much smaller and significant only for the proportion of children who became seronegative. In addition children vaccinated at age 10 to 14 years were more likely than younger children to retain a titer at least 4-fold higher than baseline at 2 years after revaccination. Discussion. We have found that EZ vaccine administered by aerosol gives better short term responses and better antibody persistence during 2 years after revaccination than EZ or SW by subcutaneous injection. The rate of decline in antibody level flattened in all vaccine groups 1 year after revaccination, suggesting that the differences in initial response might have a long term effect on offering increased protection to children who received EZ by aerosol. Children receiving vaccine by subcutaneous injection were 2 to 3 times more likely to have antibody titers below 300 mIU/ml at 2 years after revaccination and also 2 to 3 times less likely to have HI titers >1:8. Although the majority of children may not develop clinical measles on exposure to wild virus, clinical measles has been demonstrated repeatedly among children with antibody titers <125 to 200 mIU/ml. 9, 10 It has also been consistently demonstrated that individuals with antibody titers over ∼500 mIU/ml show no boosting responses on revaccination and thus may be protected from subclinical infection on revaccination. 5, 6 The lack of a response to vaccine at baseline HI titers of >1:8 seen in this study suggest that there is little or no replication of the vaccine virus, and this amount of antibody would provide protection against clinical/subclinical infection on exposure to wild measles virus in a similar way. Hence it is reasonable to predict that the higher the antibody titer, the greater is the protection from clinical disease and the lower is the chance that a child would be involved in measles transmission after exposure to wild virus. The higher antibody titers induced by aerosol may have implications for subsequent generations. Antibody titers are lower after measles vaccination than after natural measles; thus infants born to mothers with vaccine-induced immunity lose maternal antibody at an earlier age than those born to mothers who had measles disease. The administration of vaccine by aerosol to young adults, such as in mass catch-up vaccination campaigns, could therefore boost immunity of women in their early childbearing years and provide longer protection to their infants. Because countries aiming for measles elimination are changing to a schedule involving primary vaccination in the second year of life, 12, 13 this would reduce the chance that a large window of susceptibility would occur in infants born to mothers who were vaccinated in childhood. Despite ethnic differences, children receiving aerosolized vaccine responded better than children receiving vaccine by injection. This shows the aerosol to be more immunogenic than injection, irrespective of race. Age-related differences in response to revaccination usually reflect differences in antibody level before revaccination. Because our antibody assay was of low sensitivity, we may have missed differences in preexisting antibody among children who were classified as “seronegative” before revaccination. Younger children had shorter intervals since their previous dose of vaccine and thus were likely to have had higher baseline antibody titers (although below our threshold). The factors responsible for instability of nebulized SW vaccine are being investigated in ongoing studies. An unappreciated rapid loss of potency during nebulization may be a factor underlying the lesser seroresponses seen rather consistently in previous comparisons of SW and EZ aerosols. Our previous findings of strain differences in the response to revaccination by subcutaneous injection were less marked at 2 years follow-up, although serosusceptibility remained significantly more frequent in the SWsc group. Persistence of antibodies at titers that afford protection against clinical/subclinical infection makes the aerosol route of administering the measles vaccine very attractive. The main public health implication of our study is the potential to use the aerosol route for administration of vaccine in mass campaigns with associated increased protection from measles and less viral replication on exposure to measles virus in aerosol vaccinees as well as in infants of mothers vaccinated by this route. Acknowledgments. Financial support was received from the World Health Organization (Vaccines and Other Biologicals), Geneva, as well as the Medical Research Council, South Africa. We thank SmithKline Beecham Biologicals, Rixensart, Belgium, for providing vaccines used in this study. We thank Dr. David Brown, Dr. Cathy Roth, Natalie Tonkovic, Georgia Roberts and Rashpal Hunjan from the Central Public Health Laboratory Services, London, UK.

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