Immunogenicity of measles-rubella vaccine administered under India’s Universal Immunization Programme in the context of measles–rubella elimination goal: A longitudinal study

Abstract

Background & objectives: 

There is a paucity of data regarding immunogenicity of recently introduced measles–rubella (MR) vaccine in Indian children, in which the first dose is administered below one year of age. This study was undertaken to assess the immunogenicity against rubella and measles 4-6 wk after one and two doses of MR vaccine administered under India’s Universal Immunization Programme (UIP).

Methods: 

In this longitudinal study, 100 consecutive healthy infants (9-12 months) of either gender attending the immunization clinic of a tertiary care government hospital affiliated to a medical college of Delhi for the first dose of routine MR vaccination were enrolled. MR vaccine (0.5 ml, subcutaneous) was administered to the enrolled participants (1^st dose at 9-12 months and 2^nd dose at 15-24 months). On each follow up (4-6 wk post-vaccination), 2 ml of venous blood sample was collected to estimate the antibody titres against measles and rubella using quantitative ELISA kits. Seroprotection (>10 IU/ml for measles and >10 WHO U/ml for rubella) and antibody titres were evaluated after each dose.

Results: 

The seroprotection rate against rubella was 97.5 and 100 per cent and against measles was 88.7 per cent and 100 per cent 4-6 wk after the first and second doses, respectively. The mean (standard deviation) titres against rubella and measles increased significantly (P<0.001) after the second dose in comparison to the levels after the first dose by about 100 per cent and 20 per cent, respectively.

Interpretation & conclusions: 

MR vaccine administered below one year of age under the UIP resulted in seroprotection against rubella and measles in a large majority of children. Furthermore, its second dose resulted in seroprotection of all children. The current MR vaccination strategy of two doses, out of which the first is to be given to infants below one year of age, appears robust and justifiable among Indian children.

Measles and rubella are two major public health concerns accounting for a significant socio-economic burden, however, these are both vaccine preventable¹. Estimated global burden of measles cases and measles deaths was around 9.7 million and 134,200, respectively, in 2015². In India, over 2.5 million children reportedly acquire measles infection each year, whereas close to 49,000 infected children die³. Rubella, which is caused by an RNA virus usually presents with mild symptoms with up to 50 per cent of the cases being asymptomatic⁴. If rubella is acquired during the first trimester of pregnancy, it may lead to miscarriage, foetal death or congenital rubella syndrome (CRS)³. So far, there is no specific treatment for CRS, and this can only be prevented by immunization³. Approximately 139,486 rubella cases were reported to the World Health Organization (WHO) between 2007 and 2018. The annual incidence was 13.9 cases per million in 2007 and 1.7 cases per million in 2018⁵. In a survey from five sentinel sites in India (2016-2018), there were 645 suspected patients with CRS, out of which 137 (21.2%) were laboratory-confirmed CRS ⁶.

In 1978, the Expanded Programme on Immunization was launched by the Government of India⁷. In 1985, it was renamed as Universal Immunization Programme (UIP) and measles vaccine was included in the schedule the same year⁷. In India, measles vaccine was being administered to children soon after completing nine months of age considering the substantial morbidity and mortality caused by the disease and balancing the seroprotection rate with the risk of infection. Although immunization at nine months against measles increases antibody titres, the levels fall in the second year of life, thus predisposing the child to measles infection. Hence, Indian children are administered the second dose at an early age (15-24 months) rather than at school entry⁸. The second dose of measles vaccine (15-24 months) became a part of the UIP in 2010^9,10. In Delhi, the second dose of measles-containing vaccine (MCV) was given as measles–mumps–rubella (MMR) since 1999 at 15-18 months of age¹¹. In compliance with the WHO/strategic advisory group of experts on immunization recommendation about measles elimination and control of rubella/CRS by 2020, the Government of India introduced MR vaccine in UIP following a nationwide MR campaign³. Recently, the goal has been revised to ‘measles and rubella elimination by 2023’¹². Both the doses of measles vaccine provided at 9-12 and 16-24 months, respectively, were replaced by MR vaccine under routine immunization³.

Studies have shown good seroprotection after one dose of rubella vaccine, but it is unclear whether there will be any compromise in seroprotection because of interference with maternal antibodies when the vaccine is given before one year of age^13–18. The experience with administration of rubella vaccine below one year of age is limited¹⁹. Although good seroprotection rates are obtained with rubella vaccine, there are some instances of vaccine failure, which are thought to arise from neutralization of live vaccine virus by pre-existing antibodies²⁰. Seroprotection against measles before one year of age is known to be insufficient^13,14,16,18; however, it is not known whether it will be further compromised if given along with rubella vaccine as MR vaccine. Furthermore, there is inadequate literature about seroprotection after two doses of MR vaccine, when the first dose is given before one year of age. As MR vaccine has been recently included in UIP³, there is no published data yet about its efficacy in Indian settings.

This study was conducted with the objective to estimate seroprotection and antibody titres against rubella and measles 4-6 wk after each dose (first at 9-12 months and second at 15-24 months) of MR vaccine administered under the Government’s UIP in the context of MR elimination goal.

Material & Methods

This longitudinal study was conducted in the immunization clinic of the departments of Paediatrics and Microbiology, University College of Medical Sciences and associated Guru Teg Bahadur Hospital, which caters mainly to low-income urban population of Delhi, over a period of 18 months (November 2018-April 2020). A written informed consent was obtained from the parents or guardians of every study participant. An approval from the Institutional Ethics Committee–Human Research (IEC-HR) of the institute was obtained prior to the commencement of the study.

Participants: Consecutive infants aged between nine and 12 months of either gender having good health and attending the immunization clinic of the study hospital for the first dose of MR vaccine were included in the study. Children who received prolonged (>4 wk) steroid therapy, blood transfusion or immunoglobulin in the past three months, those with a history of receiving other vaccines (e.g. BCG, pentavalent and hepatitis B) concurrently with MR vaccine, those diagnosed with malignancy or immunodeficiency, infants of HIV-positive mothers, those with history of measles and those with severe acute malnutrition [weight/height <-3 standard deviation (SD) or mid-upper-arm circumference <11.5 cm or bilateral pedal oedema of nutritional origin] were excluded from the study.

Procedure/intervention: Study participants presenting to the hospital between February and July 2019 for routine vaccination were screened for enrolment after obtaining written informed consent. The first dose of MR vaccine (MR-VAC; Serum Institute of India Ltd., Pune) containing Edmonston-Zagreb strain of measles (1000 CCID50) and Wistar RA27/3 strain of rubella (1000 CCID50) supplied in lyophilized form and reconstituted with diluent supplied with the vaccine from the same manufacturer was administered at a dose of 0.5 ml subcutaneously in the right upper arm under all aseptic precautions by a trained nursing staff of the immunization clinic. The children were administered the second dose of MR vaccine at 15-24 months of age. After each vaccination, the children were observed for half an hour for any adverse events. Parents of children were requested to report for follow up between four and six weeks of receiving each dose of vaccine. At follow up, 2 ml of venous blood sample was collected in a sterile tube. Serum was separated by centrifuging the clotted sample and stored at –20°C till antibody level estimation.

The levels of anti-measles and anti-rubella antibodies were estimated using the ELISA immunoglobulin G quantitative estimation kits (measles: IMMUNOLAB GmbH, Germany, sensitivity of 97 per cent and specificity of 100 per cent; rubella: Dia.Pro Diagnostic Bioprobes SRL, Italy, sensitivity and specificity of >98%)^21,22. The test was performed and interpreted as instructed by the kit manufacturer. Antibody level of 10 U/ml served as the cut-off level for seroprotection for measles²¹, and for rubella, antibody levels >10 WHO IU/ml were considered as seroprotective²². As per the manufacturer’s recommendations, range of ±20 per cent around the cut-off is defined as a grey zone and the results between eight and 12 U/ml are to be reported as borderline for the measles antibody titre; we also reported the results by omitting the proportion of seroprotection (or its absence) contributed by these borderline values.

Sample size: In a previous study by Singh et al¹⁸ from Vellore, India, 95.6 per cent (44/46) of children had seroprotective levels against rubella after MMR vaccine administered at nine months of age. For estimating this proportion at each dose, a sample size of 65 was calculated to be sufficient with 95 per cent confidence level and five per cent absolute precision. As the follow up period was of six months, we anticipated high (one third) attrition, and planned to recruit 100 children to compensate for the loss to follow up and sample processing failures.

Statistical analysis: Descriptive statistics included a proportion for categorical variables, mean (SD) and median [interquartile range (IQR)] for antibody titres. Proportion of children who had seroprotection was compared between two doses by Chi-square test. Comparison of antibody titres after one and two doses was performed by paired t test (for mean) and non-parametric tests (for median and distribution).

Results

A total of 100 infants between the ages of nine and 12 months were recruited and vaccinated with MR vaccine. The mean (SD) age at recruitment and the first dose of MR vaccination was 10.17 (0.55) months and there was a slight male predominance (n=59, 59%). The median (IQR) weight and height of infants was 8.10 (7.92, 8.50) kg and 72 (70, 73) cm, respectively. Out of 100 infants enrolled, 98 had normal nutritional status and only two infants had moderate protein–energy malnutrition as per the WHO classification. Out of these two, one infant had weight-for-height Z score below –2 and the other had height-for-age Z score below –2. Eighty infants of the 100 recruited reported for follow up between 4-6 wk after receiving the first dose of MR vaccine. The mean (SD) number of days between the first dose of vaccine and collection of blood sample was 33 (2.26) days.

Out of these 80 infants, 70 returned for the second dose of vaccine between 15 and 24 months of age. The mean (SD) age at the time of receiving the second dose of vaccine was 16 (0.44) months. All 70 children who received the second dose of MR vaccine reported for blood collection after 4-6 wk. The mean (SD) number of days between the second dose of vaccine and collection of blood sample was 32 (1.89) days. Parents/guardians did not report any clinical feature suggestive of measles or rubella during the study in the enrolled children or their family members.

After 4-6 wk of MR vaccination, the seroprotection rate for rubella was 97.5 per cent after the first dose at 9-12 months (n=80) and 100 per cent after the second dose at 15-24 months of age (n=70). All 10 children who were lost to follow up between the first and second doses of vaccination had achieved seroprotective antibody titres against rubella after the first dose of vaccine itself with a median (IQR) of 37.69 (26.66, 77.69) WHO IU/ml. In 70 children who received both doses, the antibody titres to rubella increased significantly (P<0.001) after the second dose in comparison to the levels after the first dose [mean (SD) 116.28 (51.15) vs. 55.98 (28.3) WHO IU/ml] (Table I). The mean (SD) titre after the second dose was more than twice of the level after the first dose, with a mean difference [95% confidence interval (CI)] of 60.30 (47.45, 73.15) IU/ml between the second and first doses. Figure shows antibody titres against rubella and measles after first and second doses of MR vaccination in individual child.

Figure: Antibody titres after the first and second doses of MR vaccination in individual child. ^aWHO IU/ml for rubella and U/ml for measles. FDMT, antibody titres against measles after the first dose of vaccine; SDMT, antibody titres against measles after the second dose of vaccine; FDRT, antibody titres against rubella after the first dose of vaccine; SDRT, antibody titres against rubella after the second dose of vaccine; MR, measles-rubella

The seroprotection rate against measles was 88.7 per cent after the first dose at 9-12 months (n=80) and 100 per cent after the second dose at 15-24 months of age (n=70). There were no borderline values for measles antibody concentration after either dose of the vaccine. Eight out of 10 children who were lost to follow up between the first and second doses of vaccination had seroprotective levels of measles antibodies after the first dose with median (IQR) antibody titres being 23.71 (19.50, 25.11) U/ml in these 10 children. In 70 children who received both doses, the antibody titre after the second dose was significantly higher (P<0.001) than that after the first dose [mean (SD) 27.93 (8.10) vs. 22.23 (7.43) U/ml] (Table I). The mean (SD) titre after the second dose was about 20 per cent higher than that after the first dose, with a mean difference (95% CI) of 5.7 (3.21, 8.18) U/ml between the second and first doses. Vaccine titres against rubella and measles were compared according to the timing of sample collection after vaccination (<35 days vs. >35 days). No significant difference was observed in the immunogenicity in the two groups (Table II).

Table I: Comparison of immunogenicity against rubella and measles between the first and second doses of measles-rubella (MR) vaccine in children who received both doses (n=70)

Table II: Comparison of Immunogenicity against rubella and measles according to the timing of sample collection after vaccination

Female infants had a slightly higher seroprotection rate for both measles and rubella as compared to males after the first dose, but there were no significant gender differences in seroprotection to rubella or measles after the second dose of MR vaccine. None of the children reported any adverse event during observation period in the hospital.

Discussion

In this longitudinal study, we documented that after 4-6 wk of MR vaccination, the seroprotection rates for rubella were 97.5 and 100 per cent, respectively, after the first dose of vaccine administered before one year (9-12 months) and the second dose at 15-24 months of age. The corresponding figures for measles were 88.7 and 100 per cent, respectively. The antibody titres to rubella and measles increased significantly after the second dose in comparison to the levels after the first dose. Overall, the immunogenicity against rubella after the first dose of MR vaccine administered below one year of age was good, whereas that against measles was satisfactory and all children were seroprotected after two doses of MR vaccine as per the National immunization Schedule of Government of India.

Seroprotection is defined as an antibody threshold associated with protection from infection e.g. after a vaccination or a previous infection with a microorganism²³. In the present study, it was found that seroprotection against rubella was 97.5 per cent with MR vaccine administered before one year of age, which indicates that vaccination was effective before one year of age and even when it was given along with measles vaccine as a combined (MR) vaccine. There appears to be limited interference by maternal antibodies in the uptake of rubella vaccine before one year of age. A recent study among Thai children reported a seroprotection rate of 99 per cent to rubella with MMR vaccine administered at nine months of age¹³. Redd et al¹⁴ in Atlanta reported a seroprotection rate of 91.2 per cent to rubella with MMR vaccine at nine months of age and He et al¹⁵ in China reported a seroprotection rate of 91.3 per cent to rubella with MMR vaccine in children vaccinated at eight months of age. A study carried out by Yadav et al¹⁶ in New Delhi, India, found that the seroprotection rate with MMR vaccine at nine months was 98 per cent to rubella and Klinge et al¹⁷ in Germany reported a seroprotection rate of 97.8 per cent to rubella with MMR vaccine at nine months. Similarly, a study carried out by Singh et al¹⁸ reported 95.6 per cent seroprotection rate against rubella at nine months with MMR vaccine. MR vaccine, and not MMR vaccine as used in most of these studies, was used in our study; and similar or higher seroprotection rates were documented. A recent study by Li et al²⁴ demonstrated seroconversion against rubella in 94 per cent of infants six weeks after MR vaccine administration at eight months of age. Results from our study and the study by Li et al²⁴ indicate that the seroprotection rate against rubella with use of MR vaccine is comparable to that achieved with MMR vaccine administered before one year of age. The present study along with other studies^11–16 has shown good seroprotection even after single-dose rubella-containing vaccines (RCVs). Therefore, if high coverage is achieved, rubella elimination is possible even with one dose of RCV. However, for programmatic reasons to share cold chain capacity and because two doses of rubella vaccine are safe, the WHO has recommended implementation of a 2-dose RCV schedule with the same combined MCV vaccine for both doses⁴.

In this study, we documented a seroprotection rate of 100 per cent against rubella after two doses of MR vaccine. Hansashree et al²⁵ and Saffar et al²⁶ reported >90 per cent seroprotection after two doses of rubella-containing vaccines. A study was carried out by Wanlapakorn et al¹³ in Thai children with MMR vaccine at nine months and 2.5 yr of age and titres estimated after six months and they found the seroprotection rate against rubella to be only 78.7 per cent. The reason for such a low seroprotection rate could be that antibody levels were measured six months after the second dose in this study as compared to 4-6 wk in our study, and antibody levels may have waned in few children over this period.

In our study, we reported a seroprotection rate of 88.7 per cent against measles when MR vaccine was administered before one year of age, which compares well with earlier studies utilizing MMR vaccination below one year of age. Redd et al¹⁴ and Yadav et al¹⁶ reported seroprotection rates of 87.4 and 92 per cent, respectively, to measles with MMR vaccine at nine months of age. Singh et al¹⁸ observed that the seroconversion rate against measles at nine months of age was 92.5 per cent with MMR vaccine. Wanlapakorn et al¹³ documented seroprotection rates against measles at nine months of age to be 91.7 per cent with MMR vaccine. A study by Li et al²⁴ from China showed that measles seroconversion six weeks post-vaccination with MR vaccine at eight months of age was 99 per cent. A high rate of seroconversion in a study by Li et al²⁴ could be related to a high (99%) seronegativity before vaccination as compared to lower rates in Indian studies, suggesting lesser interference by persistence of maternal antibodies^16,24. MMR vaccine is a part of National Immunization Schedule of China since 1978. Broad coverage of population with measles vaccine may lead to vaccine-induced immunity in mothers instead of natural immunity, resulting in earlier fading of passive antibody response in their infants. Results from our study and studies utilizing MMR vaccine indicate that the seroprotection rate against measles, though lower than that against rubella, is adequate when combined measles-containing vaccines are administered before one year of age^14,16. We observed a seroprotection rate of 100 per cent against measles with two doses of MR vaccine given at 9-12 months and 15-24 months of age. Ceyhan et al²⁷ reported a seroprotection rate of 90 per cent with MMR vaccine at 15 months. Bhargava et al²⁸ reported a seroprotection rate of 70-100 per cent with MMR vaccine at 15-24 months. A 100 per cent level of protection against measles using two doses of MR vaccine obtained in our study needs to be confirmed in other settings. It is possible that by utilizing less antigens in MR vaccine as against MMR vaccine, the level of seroprotection against measles is higher because of no interference of antibody response by other antigen.

There has been some information that the first dose of measles vaccine given below one year may result in some response suppression for the second dose²⁹. In a systemic review and meta-analysis³⁰, it was found that although no effect was found on the seropositivity after MCV2, GMTs and avidity were lower after subsequent doses of MCV when the first dose was given before nine months of age³⁰. In our study, the first dose of MR was given after nine months (9-12 months) of age as per UIP. Seropositivity was not affected and seroprotection remained 100 per cent after the second dose of MR vaccine, however, the avidity of antibodies was not assessed in our study.

The main strength of this study was its longitudinal nature, which allowed us to measure antibody titres at two time points with each dose of vaccine given at two different ages. Furthermore, the study was conducted in programme mode with the MR vaccine being administered under State’s immunization programme with the use of routine healthcare staff implementing the routine immunization programme. However, a limitation of this study was follow up loss of 20 per cent for serological assessment after the first dose and another 10 per cent for the second dose. However, this was accounted for and the actual sample size enrolled was higher than the calculated sample size. So this is unlikely to have any bearing on the interpretation of results. Another limitation of our study was that we did not evaluate long-term serological response after two doses of vaccine as per the immunization programme. A recent study was conducted by Hanashree et al²⁴ on long-term seroprotection rates following the second dose of measles as MMR vaccine at 15 months in Indian children. The seropositivity rate against measles at 4-6 yr and at 9-12 yr of age declined despite two doses of measles. The seropositivity rate was 83.3 and 96.7 per cent against measles and rubella, respectively, at 9-12 yr of age. In another study conducted by Gomber et al³¹, antibody levels against measles, mumps and rubella were measured at 4-6 yr of age in children who had received one dose of measles vaccine at nine months and one dose of MMR vaccine at 15 months of age. It was found that seropositivity for measles, mumps and rubella were 20.4 per cent, 87.4 per cent and 75.7 per cent, respectively, showing poor long-term protection. In our study, we could assess only seroprotection offered by the vaccination and not the effectiveness by evaluating the impact of such a vaccination strategy on disease incidence. We did not measure antibody levels before either dose of the MR vaccine for either antigen because of ethical and feasibility issues. Furthermore, there was also no control group of unvaccinated infants. The laboratory analysis was also not carried out in duplicate; though, it is unlikely to have influenced the results because the borderline levels were not observed in our study, and minor differences on repeated measurements were unlikely to affect the seroprotection proportions.

The current MR vaccination strategy of two doses, out of which the first is given below one year of age, appears sound and justifiable in Indian infants, because it provides excellent level of seroprotection against both measles and rubella in the short term. The findings from this study may be generalizable to most settings where the first dose of MR vaccine is being administered below one year of age. However, as the study children were mostly well nourished and healthy, these results may not be applicable to children with severe malnutrition and chronic comorbidities, who may not be able to generate similar level of seroprotection. Further studies should be conducted on these vulnerable groups to assess the long-term protection offered by this strategy not only on antibody titres but also on the actual disease incidence.

Overall, in this study the MR vaccine administered below one year of age achieved seroprotection against rubella in a large majority (97.5%) of infants. The seroprotection against measles is also achieved by a large majority (88.7%) of infants. Further, with the second dose of vaccine administered at the age of 15-24 months in these infants, almost all children are likely to achieve seroprotection against rubella as well as against measles with a significant rise in titres as compared to that after first dose. The seroprotection against rubella is high even after a single dose of MR vaccine administered below one year of age and it is possible that these infants remained seroprotected even if the second dose is not administered. In resource-constrained settings, it is worthwhile to explore the use of the second dose as standalone measles vaccine rather than MR vaccine, whereas if affordable, MMR vaccine can replace MR vaccine in the immunization programme. Long-term seroprotection achieved by this strategy needs to be evaluated in future studies. The impact of such vaccination strategy on disease burden also needs to be evaluated through large-scale multicentric community-based studies.

Financial support & sponsorship: Intramural grant (MD thesis grant batch 2018-2021) from the University College of Medical Sciences, Delhi-95.

Conflicts of Interest: None.