Low seroconversion rates to measles vaccine among children in Nigeria.

Low seroconversion rates to measles vaccine among children in Nigeria* F.D. Adu,) O.A.O. Akinwolere,2 0. Tomori,3 & L.N. Uche4 The Nigerian Expanded Programme on Immunization (EPI) was assessed with particular reference to measles immunization. Of 150 children who received measles vaccine at the Institute of Child Health, University of Ibadan, Nigeria, 82 (54.7%) seroconverted. The immune response was directly related to the titre of the vaccines used. Vaccines whose titres were 10J- to 101.7 stimulated immune responses in 0-25% of vaccinees, those with titres in the range 10-21 to 10-25 stimulated responses in 12-47.6%, while those with titres of 10-27 to 10-34 stimulated responses in 87.5-100% of vaccinees. Only one of the vaccines used had a titre that met the minimum WHO required standard of log 10-3 TCID5Q at the point of vaccination. Introduction The Nigerian Expanded Programme on Immuniza- tion (EPI), which began operations in 1980, was relaunched in 1984 with huge success in terms of vaccination coverage. Since then, EPI has concentra- ted on mass vaccination efforts in an attempt to cover a large majority of the target population. What has, however, been lacking is a laboratory back-up system to ensure that the vaccines are potent on arrival in Nigeria and throughout the storage period up to the time of delivery to the vaccinee, and that the majority of the children vaccinated are actually immunized. Any successful vaccination programme must provide a highly potent vaccine, with a long expiry date, a reliable cold-chain system, and ensure that at the point of vaccination the delivery technique does not render the vaccine impotent before it is used. Recent reports of unusually large numbers of cases of measles among vaccinated children point to the urgent need for a laboratory back-up service for EPI in Nigeria. As part of a collaborative effort between the Department of Virology and the Institute of Child Health, University College Hospital, Ibadan, a pro- gramme to monitor vaccines before, during, and after use, as well as the immune responses of the vaccina- ted children, was begun in Ibadan in 1989. This article reports the initial findings. . From the College of Medicine, University of Ibadan, Ibadan, Nigeria. Senior Lecturer, Department of Virology. Requests for reprints should be sent to this author. 2 Medical Research Fellow, Institute of Child Health. 3 Professor, Department of Virology and Director, Postgraduate Institute of Medical Research and Training. 4 Technician, Department of Virology. Reprint No. 5301 Materials and methods Study population The study was carried out between April 1989 and February 1990 on children aged at least 9 months who were attending the Institute of Child Health, University College Hospital, Ibadan, for measles immunization. The children were drawn from the low, middle, and upper socioeconomic classes of the population. All the children were in a good state of health at the time of vaccination. Samples of blood were collected before vaccination by finger puncture on to filter-paper (ROPACO®), as described by Nakano (1). Each sheet of filter-paper consisted of a rectangle of 5 cm x 15.2 cm on which four circles (each 12 mm in diameter) had been imprinted. The blood from one child was used to soak each of the circles on a sheet. The name, sex, and age of each child, together with the vaccine batch and date of vaccination were marked on the filter-paper. The filter-papers were then dried at room temperature and stored at -20 °C until used. After the children had received a subcutaneous injection of 0.5 ml of measles vaccine (supplied by the EPI centre), their mothers were advised to bring them back 8 weeks later for post-vaccination evaluation. The vaccina- tions were performed by nursing sisters who had received additional training in administering EPI vaccines, thereby ensuring that a proper technique was used and that the vaccine was stored at the correct temperature in the clinic. Serum extraction and treatment were carried out as described previously (1). The final dilution of the serum was 1:10, and the accuracy of this was cross-checked using the empirical method, as described by Mattews et al. (2). Bulletin of the World Health Organization, 70 (4): 457-460 (1992) © World Health Organization 1992 457 F.D. Adu et al. Vaccine potency testing Both lyophilized and diluted measles vaccines were used in the study. Small aliquots of the diluted vac- cines were collected immediately after rehydration at the vaccination centre and stored at -70 °C until they were titrated for potency. Lyophilized, undiluted vaccines obtained from the Federal Vaccine Central Store, Oshodi, Lagos, and stored at -70 °C, were also titrated. Vaccine potency was determined accor- ding to WHO guidelines on triplicate samples of confluent monolayers of Vero cells in 24-well tissue- culture plates. The cells were incubated at 37 °C and examined daily for 10 days. The virus end-point was calculated using the method described by Reed & Muench (3). Haemagglutination inhibition test The haemagglutination inhibition test (HI) was per- formed as described by Munube (4). Serial twofold dilutions of sera were prepared from the 1:10 dilu- tion obtained from the extraction, and 4 HA (haemagglutinating) units per 0.025 ml of commer- cially available measles antigena was added to each serum dilution plate and incubated for 1 hour at 37 °C, whereupon 0.05 ml of a 1% suspension of monkey red blood cells was added. On each plate, as controls, were also run samples of red blood cells, and known positive and negative sera and antigens. The wells were incubated at 37 °C and the results were read when the reactions in the control wells were satisfactory. Vaccine stability test A few of the vaccines were tested for stability at 37 °C, 45 °C, and -70 °C; for this purpose, three ampoules of each batch of vaccine were selected and one ampoule each was stored at one of these tem- peratures. On days 1, 2, and 3 one ampoule was re- hydrated according to the manufacturer's instruc- tions and titrated in Vero cells. Results Of the 284 children, only 16 (5.6%) exhibited measles antibody (HI titre, 10-160) prior to vaccina- tion (Table 1). A total of 150 (52.8%) of the 284 children who received measles vaccine returned for post-vaccination screening, and of these, 82 (54.7%) seroconverted (titres, 10-320) following vaccination; 68 (45.3%) did not seroconvert (Table 1). Only 7 (43.8%) of the 16 children with pre-vaccination a Virion Laboratories Ltd, Ruschlikon, Switzerland. measles antibodies returned for post-vaccination screening. Five exhibited a reduced antibody titre, while two had an increased titre after vaccination (titre range, 10-40). Potency test Only one of the vaccines used in the study met the minimum WHO required standard of log i0-3TCID50. The titres of the various batches are shown in Table 2. Stability test The results indicate that the vaccines were not stable at 45 °C, since those vaccines kept at this tempera- ture for 3 days had titres that were 2 logl0 units less than at the start of the test. In contrast, vaccines kept at 37 °C or -70 °C for 3 days still retained their original titres (Table 3). Vaccine titre and immune response The relationship between measles vaccine titre and the development of measles antibody is shown in Table 2. The immune response stimulated by the dif- ferent vaccines was dependent on their titre. Four vaccine batches with titres between log 101.0 and log 10'-7 stimulated an immune response in 0-25% of the vaccinated children, while two batches with titres of log 1021 and log 102.5 stimulated an immune response in 12.5% and 47.6% of the children, respectively. Seroconversion occurred among 30 (94%) of the 32 children who received vaccines with titres of . log 102.7. Discussion Our results show that only 82 (54.7%) of the chil- dren who received measles vaccines seroconverted. Of these children, 71 (86.5%) had low-level antibody titres (1:10 to 1:40). A total of 16 children had pre-vaccination measles antibodies. These children must therefore have been exposed to the virus before 9 months of age or still exhibited circulating matemal antibodies to measles. Previous studies have reported the pres- ence of such maternal antibodies in children aged over 12 months (5, 6). In our study seven such chil- dren returned for post-vaccination screening, of whom five had become seronegative, while two had developed an anamnestic response to the vaccine. During the study only one of the vaccines used had a titre at least log l0-3 at the point of vaccina- tion. However, according to the manufacturers all the vaccines had titres in the range 104.8 to 1053. The titres were, however, not checked immediately upon receipt of the vaccines. The results of the stability 458 WHO Bulletin OMS. Vol 70 1992 Low seroconversion rates to measles vaccine in Nigeria Table 1: Results of the haemagglutination inhibition test on pre- and post-vaccination sera from children who received measles vaccine No. of Age Type of Titre range: children (months) sera No. positive No. negative <10 10 20 40 80 160 320 284 9 Pre-vaccination 16 (5.6) 268 (94.3) 268 9 2 2 2 1 - 150 9 Post-vaccination 82 (54.7) 68 (45.3) 68 26 20 25 6 4 1 a Figures in parentheses are percentages. Table 2: Titres and immune responses produced by the measles vaccines used in the study Date of Titre Immune response Name of vaccine Manufacturer Batch/lot number use (TCID50) stimulated Zagreb, Croatia Zagreb, Croatia Sclavo, Italy Osaka, Unijapan Sclavo, Italy Sclavo, Italy Sclavo, Italy Sclavo, Italy Sclavo, Italy 153 164/4 36A30 FL12/E60 36A33 36A28 33A33 33A13 33A30 7 June 1989 5 June 1989 30 August 1989 26 July 1989 6 September 1989 9 August 1989 13 September 1989 27 September 1989 20 September 1989 102.1 10-1 10-1.7 10-1 10-1 10-25 10-27 10-34 1 027 1/8 (12.5) a 0/1 (0) 0/7 (0) 0/7 (0) 1/4 (25) 10/21(47.6) 9/9 (100) 14/15 (93.3) 7/8 (87.5) Figures in parentheses are percentages. Storage tempera- Initial titre ture (OC)/day 10-3 -70/3 37/3 45/3 10-2 -70/3 37/3 45/3 1025 -70/3 37/3 45/3 n selected Meanwhile, we suggest that the cold chain should be evaluated and that the storage conditions of the vac- cines should be examined. Also the use of highly Final titre heat-stable vaccines should be encouraged, vaccines should be randomly tested before use, and any batch10 3 with a titre of <10-3 should be discarded.10-3 10 10-2 10-2 10-1 10-25 10-24 10-1 Acknowledgements This study was supported by UNICEF. The technical assistance of Mr S.J. lkpeme and the secretarial work of both Miss S.N. Anyanwu and Miss 0. Afelumo are grate- fully acknowledged. Resume test showed that the vaccines were relatively stable at 37 °C and -70 °C but not at 45 'C. The vaccina- tions were performed in the clinic, where the aver- age room temperature was normally air-conditioned to below 30 'C, and the vials of vaccines were usually stored on ice packs provided by EPI. The immune response stimulated by each vac- cine was directly dependent on its titre (Table 2). The poor response of the vaccinees in this study is attributable to the low vaccine titres. Several reasons could have been responsible and further studies are being carried out to identify the factors that may have led to these subpotent vaccine titres. Faibles taux de seroconversion apres vaccination antirougeoleuse chez l'enfant au Nigeria Le programme elargi de vaccination (PEV) du Nig6ria a ete 6valu6 en ce qui concerne la vacci- nation antirougeoleuse. Sur 150 enfants ayant recu le vaccin antirougeoleux a lInstitut de P6dia- trie de l'Universit6 d'Ibadan (Nigeria), 82 (54,7%) ont pr6sente une s6roconversion. La reponse immunitaire etait directement liee au titre du vac- cin utilis6. Les vaccins dont le titre etait de 10-1-101,7 (DICT50) ont suscite une reponse WHO Bulletin OMS. Vol 70 1992 Conpharma Conpharma Morbilvax Bikem Cam. Morbilvax Morbilvax Morbilvax Morbilvax Morbilvax Table 3: Results of the stability tests o batches of measles vaccine Vaccine batch 30A30 36A30 CXV (115) 459 F.D. Adu et al. immunitaire chez 0 a 25% des vaccines, ceux dont le titre etait compris entre 1 o-2 1 et 1 o-25 chez 47,6% des vaccin6s, et ceux dont les titres etaient compris entre 1 0-27 et 10-3'4 chez 87,5 a 100% des vaccin6s. Seul un des vaccins utilises avait un titre satisfaisant a la norme minimale requise par l'OMS, soit log 1 0-3 DICT50 au point d'injection. Dans cette etude, la m6diocrit6 de la reponse a la vaccination est imputable aux faibles titres des vaccins. Ces faibles titres peuvent avoir plu- sieurs origines, que l'on cherche actuellement a identifier. En attendant, nous proposons que la chaine du froid soit 6valu6e et que les conditions de conservation des vaccins soient examinees. II faudrait 6galement encourager l'emploi de vac- cins de bonne thermostabilite, tester des 6chan- tillons de vaccins pris au hasard avant leur utili- sation, et rejeter tout lot ayant un titre inferieur a log 10-3 DICT50. References 1. Nakano, J.H. et al. Microtitre determination of measles haemagglutination inhibition antibody with filter-papers. Journal of clinical microbiology, 17: 860-863 (1983). 2. Mattews, H.M. Parasitic diseases: testing with filter- paper blood spots. Laboratory management, 9: 55-62 (1981). 3. Reed, L.J. & Muench, H. A simple method of esti- mating fifty percent end-points. American journal of hygiene, 27: 493-497 (1938). 4. Munube, G.M.R. Measles seroimmunity in rural non-vaccinated children of Busoga District, Uganda. East African medical journal, 56: 335-338 (1979). 5. Albrecht, P. et al. Persistence of maternal antibody in infants beyond 12 months: mechanism of measles vaccine failure. Journal of pediatrics, 91: 715-718 (1977). 6. Abdurrahman, M.B. et al. Measles antibody levels from birth to 9 months of age in Nigerian infants. African journal of medical sciences, 11: 113-115 (1982). 460 WHO Bulletin OMS. Vol 70 1992