Archives ofDisease in Childhood 1996;75:298-303

Combined diphtheria, tetanus, pertussis, andHaemophilus influenzae type b vaccines for primaryimmunisation

Frank Bell, Adele Martin, Christine Blondeau, Carol Thornton, Janet Chaplais,Adam Finn

AbstractA total of 146 infants were immunised atages 2, 3, and 4 months with a combineddiphtheria, tetanus, pertussis (DTP) -Haemophilus influenzae type b (Hib)tetanus toxoid conjugate (PRP-T) vaccine(Pasteur Merieux) to assess the antibodyresponse and adverse events associatedwith immunisation. Adverse events, in-cluding fever, were recorded by parents ina diary for three days following eachinjection. Blood was taken before the firstimmunisation and four weeks after thethird immunisation to assess antibodyresponse. Data were compared with thosefrom historical controls who had receivedDTP and PRP-T vaccines by separateinjection. The combined vaccine was welltolerated. Rates of local and general reac-tions were similar to those reported forinfants immunised by separate injection.All infants achieved protective antibodytitres (> 0.01 IU/ml) for diphtheria andtetanus; 98% acquired Hib (PRP) anti-body > 0.15 ttg/ml and 82.5% > 1.0 tg/ml.Pertussis antibody titres (pertussis toxin,filamentous haemagglutinin, total aggluti-nins, and agglutinins 2 and 3) showedappreciable rise following immunisation.DTP and PRP-T vaccines provide similarantibody responses and adverse effectswhether mixed in the same syringe oradministered by separate injection. Thevaccines could be combined for use in theUnited Kingdom primary immunisationschedule.(Arch Dis Child 1996;75:298-303)

Keywords: diphtheria tetanus pertussis vaccine, Haemo-philus influenzae type b vaccine, immunisation.

The introduction of Haemophilus influenzaetype b (Hib) immunisation to the universalschedule in the United Kingdom in 1992 hasled to a dramatic reduction in reported rates ofinvasive haemophilus disease in children.'However, extending the protection offered byimmunisation doubled the number of injec-tions given to infants in the primary scheduleas Hib and diphtheria, tetanus, and pertussis(DTP) vaccines have been given separately ateach immunisation visit. Combining the in-jected antigens of the primary schedule into asingle injection would reduce discomfort forinfants, so might improve parental acceptabil-

ity and vaccine uptake. In the United Kingdoma combined DTP-Hib vaccine would reducethe number of immunising injections by twomillion each year. Healthcare staff wouldwelcome the time saved by carrying out thesimpler procedure. Demonstrating the safetyand immunogenicity of vaccine combinationswill become increasingly important as newantigens are introduced into the universalprimary immunisation schedule.

Several conjugate vaccines derived from Hibcapsular polysaccharide have been developedand considered for primary immunisation ofinfants. Previous studies, using polyribosylribi-tol phosphate conjugated with tetanus toxoid(PRP-T) given in the same syringe as DTP,have shown the combination to be welltolerated.2 However, some reports have sug-gested a reduction in vaccine immunogenic-ity,67 particularly in the response to pertussisagglutinins.8-0 It is unclear how much variationis attributable to the mixing of antigens andstill more difficult to assess the clinical signifi-cance of minor changes in mean antibody titre.A previous study mixing these vaccines identi-fied lower levels of PRP antibody in the groupreceiving the combined vaccine.6 Mean titres inthe mixed group were still several times higherthan the level thought to confer long term pro-tection, and all infants achieved levels abovethis threshold. Protective thresholds have notbeen established for pertussis, which makesinterpreting differences in immune responsemore difficult.

Concerns about impaired antibody responsemay be particularly relevant in the UnitedKingdom in view of the earlier completion ofthe accelerated schedule at 4 months, whichmay itself interfere with optimal immune

11 12response, and in the absence of boosterdoses of pertussis or Hib vaccines beyond thisage.We sought to establish whether combined

administration of DTP with PRP-T (a Hibconjugate vaccine in current use in the UnitedKingdom) would provide satisfactory protec-tion for infants with an acceptable profile ofadverse events. Accepted protective antibodythresholds exist for Hib,"-16 diphtheria, andtetanus,'7-'9 allowing comparisons to be madewith previous studies of infants immunisedin the United Kingdom acceleratedschedule." 12 2022We immunised infants with a vaccine

combining DTP and Hib (PRP-T) mixed in a

UniversityDepartment ofPaediatrics, SheffieldChildren's Hospital,SheffieldF BellA MartinA Finn

Pasteur MerieuxLaboratoire deSero-ImmunologieClinique, Val de Reuil,FranceC Blondeau

Centre for AppliedMicrobiology andResearch, PortonDown, WiltshireC Thornton

Department of ChildHealth, CommunityHealth SheffieldJ Chaplais

Correspondence to:Dr F Bell, UniversityDeparunent of Paediatrics,Sheffield Children's Hospital,Western Bank, Sheffield S102TH.

Accepted 3 July 1996

298

on 12 August 2019 by guest. P

rotected by copyright.http://adc.bm

j.com/

Arch D

is Child: first published as 10.1136/adc.75.4.298 on 1 O

ctober 1996. Dow

nloaded from

Vaccines for primary immunisation

'bypass' syringe given at age 8, 12, and 16weeks, assessed the frequency and severity ofadverse events and examined the antibodyresponse to immunisation.

MethodsParents of healthy infants were approached onthe postnatal wards of the two obstetric unitsserving the city of Sheffield, England and giveninformation about the study. Families whoexpressed interest by returning a reply slipwere seen later at home, where writteninformed consent for the study was obtained.Babies born at less than 37 weeks' gestation orweighing less than 2500 g at birth wereexcluded. Infants of families taking part werescheduled to receive immunisations at home atage 8, 12, and 16 weeks. All received a singleinjection at each visit combining Hib-tetanustoxoid conjugate (PRP-T) and adsorbed diph-theria, tetanus, and whole cell pertussis vac-cines (Pasteur Merieux). This was given byintramuscular injection in the right antero-lateral thigh using a 25G needle. Antigens weremixed immediately before administrationusing a 'bypass' syringe which allows liquidDTP vaccine in the upper barrel to reconsti-tute dried PRP-T in the lower portion of thesyringe, resulting in an injected volume of 0.5ml. Single batches of PRP-T and DTPvaccines were supplied by the manufacturer forthe study. Oral polio vaccine (Wellcome) wasgiven at each immunisation visit.

Parents were given a surface activated liquidcrystal thermometer (Body-temp, TommeeTippee, Cramlington, UK) and a diary inwhich to record temperature twice daily, withdetails of other adverse events for the threedays following immunisation. They were askedto note redness or swelling at the injection site(greater than 2.5 cm diameter), to assess infantcrying, activity, and feeding, and to record anyuse of paracetamol. Parents were advised togive paracetamol if their baby seemed to be indiscomfort or had a temperature of 38°C orabove. We contacted parents by telephone atone and three days following immunisation toask about unwanted effects and to confirmdiary information.Venous blood was taken before the first

injection and again four weeks after the thirdimmunisation to assess antibody responses. Alocal anaesthetic cream (EMLA, Astra Phar-maceuticals, Kings Langley, UK) was appliedbefore blood sampling at 5 months. Sera wereanalysed for antibody to diphtheria and tetanustoxoids, PRP, and pertussis antigens (pertussistoxin (PT), filamentous haemagglutinin (FHA),total agglutinins and agglutinins 2 and 3 (fimbrialantigens)). Antibody to PRP was measured byradioimmunoassay. Diphtheria and tetanus anti-toxins, antibodies to PT, FHA, and agglutinins 2and 3 were measured by enzyme linked immuno-sorbent assay (EHISA). Total pertussis aggluti-nins were determined by serial dilution of serumuntil precipitation with pertussis suspension wasno longer observed.

All assays except for pertussis agglutinins 2and 3 were performed at Val de Reuil, France(laboratory 1). There are no universally

accepted standards or methods for pertussisantigen assays, so postimmunisation sera wereadditionally assayed (ELISA)23 for pertussistoxin and FHA at the Centre for AppliedMicrobiology and Research, Porton Down(laboratory 2) to allow comparison with previ-ous reports from the United Kingdom. Thislaboratory was also responsible for preimmuni-sation and postimmunisation assays for agglu-tinins 2 and 3.23 Where results lay below thelimit of detection for the assay, a value of onehalf the lower limit was used to calculategeometric mean titres.

STATISTICSAntibody levels following immunisation werecompared in groups of infants with low andhigh preimmunisation titres by the Mann-Whitney U test for non-parametric data.Analyses were performed using SPSS for Win-dows Release 6.0 (SPSS, Chicago, Illinois,USA). Categorical data were compared usingthe X2 test or Fisher's exact test where fewerthan five infants were expected in any group.Approval for the study was obtained from theNorth and South Sheffield research ethicscommittees.

ResultsOne hundred and forty six infants (71 girls)were recruited to the study. Twenty four percent of the parents approached agreed to takepart. All immunisations and blood tests tookplace on or within two weeks of the dates whenthese were due. Median ages at immunisationand at final blood sampling were 8.1, 12.4,16.6, and 20.9 weeks. Two infants did notcomplete the study: temporary loss of contactwith one family led to a delay between secondand third immunisations, and a second familymoved away before the final blood sample wastaken.One infant developed swelling involving the

major part of the thigh following secondimmunisation, precluding administration of athird dose of the combined vaccine. Sheremained otherwise well; swelling and ery-thema had resolved by four days. Immunisa-tion was completed with DT, Hib, and poliovaccines and this infant excluded from sero-logical analysis. No other adverse effects metDepartment of Health criteria for severereactions to immunisation.24 A minority ofinfants developed appreciable erythema orswelling at the immunisation site (table 1).Modest rates were reported for prolonged cry-ing and for disturbed feeding (table 1). Up toone half of all infants were febrile at some timein the three days following immunisation.Temperatures > 38°C were recorded lessfrequently with succeeding doses (table 1).Serum was available from 143 infants

following immunisation with three doses ofcombined vaccine. Insufficient blood wasobtained from one infant for preimmunisationanalysis. All achieved protective antibody titres(over 0.01 IU/ml) for diphtheria and tetanus(table 2). Three infants failed to achieveprotective titres for PRP (table 2). Geometricmean antibody titres for these antigens before

299

on 12 August 2019 by guest. P

rotected by copyright.http://adc.bm

j.com/

Arch D

is Child: first published as 10.1136/adc.75.4.298 on 1 O

ctober 1996. Dow

nloaded from

30Bell, Martin, Blondeau, Thornton, Chaplais, Finn

Table 1 Number (percentage) of infants experiencingadverse events in the 3 days following immunisation(n=140-146)

Dose

1 2 3

Erythema > 2.5 cm 6 (4) 12 (8) 9 (6)Swelling > 2.5 cm 16 (11) 15 (10) 13 (9)Crying> I hour 16 (11) 9 (6) 14 (10)Inconsolable > 4 hours 0 (0) 2 (1) 1 (1)

Disturbed feedingFed less than usual 59 (41) 30 (21) 33 (23)Missed feeds 17 (12) 6 (4) 10 (7)

Fever > 38°COn first evening 52 (36) 26 (18) 17 (12)Any (over 3 days) 77 (53) 44 (31) 35 (24)

and after immunisation are shown in table 3.There were marked increases in titres ofantibody to the four pertussis antigens meas-

ured (table 4).The influence of maternally acquired anti-

body was assessed by comparing antibodytitres before and after immunisation. Infantswith titres above protective threshold fortetanus and diphtheria before immunisationachieved lower titres at 5 months than thosewith initial levels of antibody below these titres(table 5). This was more clearly seen fortetanus, 38 infants (27%) experiencing a fall intitre between 2 and 5 months of age despiteimmunisation. Similarly, infants with low anti-body levels before immunisation achievedhigher titres for pertussis toxin, FHA, andagglutinins 2 and 3. No effect of preimmunisa-tion titre on antibody levels at 5 months was

shown for PRP or for total pertussis aggluti-nins.

DiscussionCombining the antigens administered by sepa-rate injections at one visit would be welcomedby infants, parents, health workers, and admin-istrators. As new vaccines are proposed forinclusion in the primary immunisation sched-ule, it will continue to be necessary to demon-strate the safety and effectiveness of vaccinecombinations in order to maximise protectionagainst infectious diseases without an unac-

ceptable increase in immunisation visits or

injections.Several studies have addressed the immuno-

genicity and tolerance of a number of DTPvaccines mixed with PRP-T in other settings,given by more extended schedules.2'-0 Thosethat have considered PRP-T in the UnitedKingdom schedule, either combined or as a

separate injection, have used the Wellcome(Evans) DTP vaccine.20 22 Certain DTP/Hibconjugate vaccine combinations have beenlicensed, are in use in the USA,25 and haverecently been recommended in the UnitedKingdom.26There was no obvious bias in maternal age,

race, social class, or geographical distributionwithin the city in the sample recruited to thisstudy. The combined PRP-T/DTP vaccinegiven at 2, 3, and 4 months was well toleratedby these infants. Only one child failed to com-plete immunisation for all antigens because ofa local reaction following the second dose of

the combined vaccine. Rates of redness (4-8%)and swelling (9-11%) at the injection site weresimilar to those reported for PRP-T combinedwith Wellcome DTP22 and lower than thosereported for DTP when given either in a sepa-rate limb from PRP-T (redness 12-27%, swell-ing 12-24%),22 or given alone without Hib vac-cine (redness 10-26%, swelling 10-32%).21 27No serious systemic reactions were encoun-

tered, although over 50% of infants had fever¢ 38°C occurring at any time during the threedays after the first dose of vaccine comparedwith the low rates (3-10%) reported previ-ously.22 However, direct comparisons are diffi-cult as the methods for analysis of temperaturedata in that study are unclear. In another study,6% of infants receiving vaccines as twoseparate injections had fever after the first dosewhen parents measured temperatures oncedaily.20 Rates of fever for subsequent doses inour study (31% following the second and 24%after the third) compare with reports of 16%for both doses for combined vaccine recipi-ents22 or 5-16% and 6-24% respectively, forinfants receiving vaccine by separateinjection.20-22Our results may have been influenced by the

use of a forehead thermometer reading to thenearest whole degree (allowing interpolation tonearest 0.5'C) which may have overestimatedthe number of infants with temperature above38°C as temperatures were rounded up. Oneprevious study has reported a higher rate offever in infants receiving one of two batches ofPRP-T combined with DTP,'0 but no other hasshown an increase in febrile responses amongcombined vaccine recipients, and an earlierUnited Kingdom study found similar rates inhistorical controls given vaccine by separateinjection.22 Differences in response might alsobe explained by the use ofDTP from differentmanufacturers,28 or by greater use of paraceta-mol in other studies. Paracetamol was given byparents after 55% of immunisations in thisstudy, but these data have not been recorded inprevious reports. None of the febrile infants inthis study were seriously unwell, and no childrequired admission to hospital or interventionother than with simple antipyretics.

Several studies have addressed the immuno-genicity of mixed PRP-T/DTP in more ex-tended schedules. Some have identified modestreductions in antibody titre to PRp,6 tetanus,79and pertussis,>'0 while others have found noevidence for reduced immunogenicity.2 Onestudy showed an enhanced response to diph-theria for combined vaccine recipients,3 and inanother, infants immunised on a 2, 3, 4 monthschedule showed higher antibody titres to teta-nus.22

Infants in our study acquired satisfactoryantibody titres for each vaccine constituent. Allachieved diphtheria and tetanus titres aboveprotective threshold. Geometric mean titres(GMT) for diphtheria and tetanus were similarto those reported for United Kingdom infantsimmunised with DTP in the acceleratedschedule (table 3)1.12 21 22The mean titre for PRP in this group of

infants is similar to the figure reported by Begg

300

on 12 August 2019 by guest. P

rotected by copyright.http://adc.bm

j.com/

Arch D

is Child: first published as 10.1136/adc.75.4.298 on 1 O

ctober 1996. Dow

nloaded from

Vaccines forprimary immunisation

Table 2 Number (percentage) of infants with antibody levels above protective thresholdcompared with those reported previously for infants receiving vaccines by separateinjections1 20

PostimmunisationPreimmunisation Postimmunisation PRP-Tand DTP*PRP-TIDTP combined PRP-TIDTP combined separate sites"20

Diphtheria> 0.01 IU/mi 46/134 (34) 143/143 (100) t (100)> 0.1IU/ml 7/134 (5) 142/143 (99)

Tetanus> 0.01 IU/mi 133/142 (94) 143/143 (100) t (100)> 0.1 IU/mi 109/142 (77) 143/143 (100) 89/91 (98)

PRP> 0.15 pg/mi 43/142 (30) 140/143 (98) 105/107 (98)> 0.1 jg/ml 1/142 (1) 118/143 (82) 97/107 (91)

* Wellcome (Evans) DTP.t Numbers not given in report.DTP = absorbed diphtheria, tetanus, pertussis vaccine; PRP-T = polyribosylribitol phosphatetetanus toxoid conjugate vaccine; PRP = polyribosylribitol phosphate.

Table 3 Geometric mean antibody titres (95% confidence limits for mean) for diphtheria,tetanus, and polyribosylribitol phosphate (PRP), compared with those reported previouslyfor infants receiving vaccines by separate injectons

Preimmunisation Postimmunisation Postimmunisation PRP-T andPRP-TIDTP combined PRP-TIDTP combined DTP** separate sites" 20(n=142) (n=143) (n=103)

Diphtheria 0.007* (0.006 to 1.07 (0.95 to 1.21) 3.87(IU/ml) 0.009)

Tetanus 0.25 (0.19 to 0.33) 1.01 (0.87 to 1.16) 0.70(IU/ml)

PRP (pLg/ml) 0.12 (0.11 to 0.14) 3.65 (2.92 to 4.56) 5.01

* 134 samples available for preimmunisation diphtheria serology.** Wellcome (Evans) DTP.DTP = absorbed diphtheria, tetanus, pertussis vaccine; PRP-T = polyribosylribitol phosphatetetanus toxoid conjugate vaccine; PRP = polyribosylribitol phosphate.

Table 4 Geometric mean antibody titres (95% confidence limits for mean) for pertussisantigens compared with those reported previously for infants receiving vaccines by separateinjectwns

Preimmunisation Postimmunisation Postimmunisation PostimmunisationPRP-TIDTP PRP-TIDTP PRP-TIDTP PRP-T and DTP**combined combined combined separate siteslaboratory I laboratory lt laboratory 2t laboratory 2"1(n=142) (n=143) (n=143) (n=102)

Pertussis toxin 1.2 (1.0 to 1.5) 41 (31 to 55) 6700 (5250 to 850(EIAU/ml) 8540)

FHA (EIAU/mil) 5.7 (4.7 to 7.0) 40 (35 to 47) 6110 (5270 to 62007080)

Total agglutinins 20 (16 to 24) 420 (360 to - -

(reciprocal of 480)dilution)

Agglutinins 2 and 195* (140 to - 33 490* (26 610 38 9003 (reciprocal of 270) to 42 150)dilution)

* Preimmunisation and postimmunisation serology for agglutinins 2 and 3 performed in labora-tory 2.** Weilcome (Evans) DTP.t Different assays used for antibody to pertussis toxin and FHA in laboratories 1 and 2. Resultscannot be compared between laboratories.DTP = absorbed diphtheria, tetanus, pertussis vaccine; PRP-T = polyribosylribitol phosphatetetanus toxoid conjugate vaccine; FHA = filamentous haemagglutinin; EIAU = enzymeimmunoassay (that is, arbitrary) units.

et al from Gloucester, mixing PRP-T withDTP.22 Both are a little lower than titresreported for PRP-T administered in a separatelimb: infants in our study achieved a mean titreof 3.65 jg/ml compared with 3.4 jg/ml for thecombined vaccine in Gloucester.22 Infantsgiven vaccine by separate injection achievedGMT of 4.5 j.g/ml in that study and 5.0 jig/mlin the first United Kingdom study of thePRP-T vaccine in the Oxford region.20 Threeinfants in our study failed to achieve minimumprotective antibody titres four weeks after theprimary course was complete. The sameproportion (2%) remained unprotected inOxford,20 an expected response to Hib conju-

gate vaccines in infancy. Follow up from a largetrial of the earlier Hib polysaccharide (uncon-jugated) vaccine found that PRP titres above1 jig/ml three weeks after immunisation wereassociated with continued protection againstinvasive Hib disease in an immunisedpopulation.' '6 Eighty two per cent of ourinfants achieved titres above 1.0 ig/ml, while91% given vaccine by separate injectionsreached this threshold in Oxford.20 Theproportion achieving this titre in our study-although lower than that reported fromOxford-is consistent with experience in in-dustrialised countries of administering PRP-Tby separate injection.29 Two studies have shownequivalent GMT for PRP following immunisa-tion with DTP and PRP-T vaccines whencombined or given at separate sites51'; severalothers have found lower mean values in thecombined vaccine group,922 although this onlyachieved statistical significance in one studyfrom Chile.6

Infants completing the current study withPRP titres below 0.15 jig/ml were offered anadditional dose of vaccine at 8-10 months ofage. All three subsequently achieved antibodylevels above 1 jig/ml, suggesting effective prim-ing by earlier immunsation.There were substantial rises in antibody titre

to each of the pertussis antigens measured.Mean titres following immunisation werehigher (pertussis toxin) or close (FHA, aggluti-nins 2 and 3) to figures from previous reportsfrom the accelerated schedule (table 4).". 21 22Those studies used a different DTP vaccine;the response to pertussis antigens, particularlypertussis toxin, may vary between vaccinesfrom different manufacturers.'03'The 'accelerated' United Kingdom primary

schedule is unusual in beginning immunisationat age 2 months with completion by 4 monthsin the absence of subsequent reinforcing dosesof pertussis or Hib vaccines. The timing of theschedule has raised concerns about interfer-ence from maternal antibody still present at thetime of first immunisation." We found no evi-dence for substantial interference in this popu-lation, although infants with high levels ofmaternal antibody to tetanus and diphtheriaachieved lower titres at 5 months than thosewith lower initial levels of antibody (table 5).The clinical significance of this inhibition isuncertain. In the short term at least, all infantsachieved levels of tetanus and diphtheriaantitoxin held to confer protection againstdisease.'7"'" The effect on response to diphthe-ria was less marked, with preimmunisationtitres much lower than for tetanus, reflectingmaternal immunisation experience. Until re-cently the last dose of diphtheria toxoid hasbeen given in the United Kingdom at age 4years (school entry). Since 1994, combinedtetanus/diphtheria immunisation has been rec-ommended for school leavers. As diphtheriaantibody levels rise in the young adult popula-tion, interference with the response to primaryimmunisation may become more important.

Inhibition from passive antibody has previ-ously been demonstrated in the acceleratedschedule for tetanus, pertussis toxin, and for

3.01

on 12 August 2019 by guest. P

rotected by copyright.http://adc.bm

j.com/

Arch D

is Child: first published as 10.1136/adc.75.4.298 on 1 O

ctober 1996. Dow

nloaded from

302 Bell, Martin, Blondeau, Thornton, Chaplais, Finn

Table 5 Effect ofpreimmunisation titres on subsequent antibody levels. Infants weredivided into two groups (above and below threshold indicated) on the basis of initialantibody titre. Postimmunisation titres for these groups were compared by Mann-Whitneytest

Threshold n 1, n high Mean titre . Mean titre highpreimmunisation pre- pre- pre- pre- p Value

Diphtheria 0.01 IU/mi 85 49 1.25 0.83 < 0.01Tetanus 0.1IU/ml 33 109 1.62 0.88 < 0.001PRP 0.15 jg/ml 99 43 3.52 3.91 0.68Pertussis toxin* Median 71 71 91.5 18.0 < 0.001FHA* Median 71 71 50.3 32.5 < 0.01Total < 10/> 20 62 80 438 408 0.91

agglutininsAgglutinins 2 Median 71 71 48 591 22 504 < 0.01and 3

* Assays performed in laboratory 1.n I, pre- = number of infants with initial antibody below threshold indicated; nhigh pe- = numberof infants with initial antibody above threshold indicated; mean titre ,,w pre- = final geometricmean antibody titre in group with 'low' initial titres; mean titre high pre- = final geometric meanantibody titre in group with 'high' initial titres; PRP = polyribosylribitol phosphate; FHA = fila-mentous haemagglutinin.

pertussis agglutinins 2 and 3.11 High initialtitres of antibody to tetanus, the PRP-T carrierprotein, have also been associated with lowerantibody response to PRP in infants immu-nised with two doses of PRP-T."2 In contrast,we found higher PRP responses in infants withpreimmunisation tetanus antibody above 0.1IU/ml (p < 0.00 1, Mann-Whitney test). Theseinfants were more likely to achieve protectivethresholds for PRP; three infants with low ini-tial tetanus titres failed to reach 0.15 jg/ml forPRP, while no infant with higher initial titresremained unprotected (p = 0.01, Fisher's exacttest). Similarly, 61% of those with lowpreimmunisation tetanus titres achieved PRPantibody of 1.0 jg/ml or more, compared with89% for those with high initial tetanus titresp < 0.001, x2 test).

Factors influencing the antibody response tothe primary series-such as age at immunisa-tion, the mixing of antigens, or the nature ofthe conjugate vaccine carrier protein-may allhave implications for continued vaccine pro-tection. Concerns about impaired antibodyresponse to vaccines may increase as new anti-gens, for example acellular pertussis vaccines,are included and mixed in the primaryschedule. This may require reappraisal of theneed for booster doses of vaccine in the secondyear. We intend to investigate the persistence ofprotective antibody in a study measuring anti-body levels at age 13 months and at schoolentry.

Despite these observations, immunisation inthe manner described conferred high levels ofprotection with few adverse effects. Combiningthe injected antigens ofthe primary schedule inthe same syringe provides similar antibodyresponses to those achieved by administeringDTP and Hib (PRP-T) vaccines by separateinjection. The combined vaccine improvesacceptability and provides a basis for includingnew immunising antigens in the future.

We are grateful to Dr Robert Booy for helpful discussion inpreparation for this study. The study was supported by a grantfrom Pasteur Merieux MSD.

I Hargreaves RM, Slack MPE, Howard AJ, Anderson E,Ramsay ME. Changing patterns of invasive Haemophilusinfluenzae disease in England and Wales after introductionof the Hib vaccination programme. BMJ 1996;312:160-1.

2 Watemberg N, Dagan R, Arbelli Y, et al. Safety and immu-nogenicity of Haemophilus type b-tetanus protein conju-gate vaccine, mixed in the same syringe with diphtheria-tetanus-pertussis vaccine in young infants. Pediatr Infect DisJ 1991;10:758- 63.

3 Avendano A, Ferreccio C, Lagos R, et al. Haemophilusinfluenzae type b polysaccharide-tetanus protein conjugatevaccine does not depress serologic responses to diphtheria,tetanus or pertussis antigens when coadministered in thesame syringe with diphtheria-tetanus-pertussis vaccine attwo, four and six months of age. Pediatr Infect Dis J1993;12:638-43.

4 Kaplan SL, Lauer BA, Ward MA, et al. Immunogenicity andsafety of Haemophilus influenzae type b-tetanus proteinconjugate vaccine alone or mixed with diphtheria-tetanus-pertussis vaccine in infants. J Pediatr 1994;124:323-7.

5 Miller MA, Meschievitz CK, Ballanco GA, Daum RS.Safety and immunogenicity of PRP-T combined withDTP: excretion of capsular polysaccharide and antibodyresponse in the immediate post-vaccination period. Pediat-rics 1995;95:522-7.

6 Ferreccio C, Clemens J, Avendano A, et al. The clinical andimmunologic response of Chilean infants to Haemophilusinfluenzae type b polysaccharide-tetanus protein conjugatevaccine coadministered in the same syringe withdiphtheria-tetanus toxoids pertussis vaccine at two, fourand six months of age. Pediatr Infect DisY 1991;10:764- 71.

7 Dagan R, Botujansky C, Watemberg N, et al. Safety andimmunogenicity in young infants of Haemophilus b-tetanusprotein conjugate vaccine, mixed in the same syringe withdiphtheria-tetanus-pertussis-enhanced inactivated poliovirusvaccine. Pediatr Infect DisJY 1994;13:356-62.

8 Clemens JD, Ferreccio C, Levine MM, et al. Impact ofHae-mophilus influenzae type b polysaccharide-tetanus proteinconjugate vaccine on responses to concurrently adminis-tered diphtheria-tetanus-pertussis vaccine. JAMA 1992;267:673-8.

9 Gold R, Scheifele D, Barreto L, et al. Safety andimmunogenicity of Haemophilus influenzae vaccine (teta-nus toxoid conjugate) administered concurrently orcombined with diphtheria and tetanus toxoids, pertussisvaccine and inactivated poliomyelitis vaccine to healthyinfants at two, four and six months of age. Pediatr Infect Disy 1994;13:348-55.

10 Scheifele D, Barreto L, Meeltson W, Guasparini R, FriesenB. Can Haemophilus influenzae type b-tetanus toxoid con-jugate vaccine be combined with diphtheria toxoid pertus-sis vaccine-tetanus toxoid? Can Med Assoc Y 1995;149:1105-12.

11 Booy R, Aitken SJ, Taylor S, et al. Immunogenicity of com-bined diphtheria, tetanus, and pertussis vaccine given at 2,3, and 4 months versus 3, 5, and 9 months of age. Lancet1992;339:507-10.

12 Ramsay ME, Rao M, Begg NT, Redhead K, Attwell AM.Antibody response to accelerated immunisation with diph-theria, tetanus, pertussis vaccine. Lancet 1993;342:203-5.

13 Robbins JB, Parke JC, Schneerson R, Whisnant JK. Quanti-tative measurement of "natural" and immunization-induced Haemophilus influenzae type b capsular polysac-charide antibodies. Pediatr Res 1973;7:103-10.

14 Kayhty H, Peltola H, Karanko V, Makela PH. Theprotective level of serum antibodies to the capsularpolysaccharide of Haemophilus influenzae type b. Y InfectDis 1983;147:1100.

15 Peltola H, Kayhty H, Virtanen M, Makela PH. PreventionofHemophilus influenzae type b bacteremic infections withthe capsular polysaccharide vaccine. N Engl Y Med1984;310:1561-6.

16 Anderson P. The protective level of serum antibodies to thecapsular polysaccharide of Haemophilus influenzae type b.YrInfect Dis 1984;149:1034-5.

17 Ipsen J. Circulating antitoxin at the onset of diphtheria in425 patients. JImmunol 1946;54:325-47.

18 McComb JA. The prophylactic dose ofhomologous tetanusantitoxin. N EnglJ Med 1964;270:175-8.

19 Smith JW. Diphtheria and tetanus toxoids. Br Med Bull1969;25: 177-82.

20 Booy R, Taylor SA, Dobson SR, et al. Immunogenicity andsafety of PRP-T conjugate vaccine given according to theBritish accelerated immunisation schedule. Arch Dis Child1992;67:475-8.

21 Ramsay ME, Miller E, Ashworth LA, Coleman TJ, Rush M,Waight PA. Adverse events and antibody response to accel-erated immunisation in term and preterm infants. Arch DisChild 1995;72:230-2.

22 Begg NT, Miller E, Fairley CK, et al. Antibody responsesand symptoms after DTP and either tetanus or diphtheriaHaemophilus influenzae type B conjugate vaccines givenfor primary immunisation by separate or miixed injection.Vaccine 1995;13:1547- 50.

23 Rutter DA, Ashworth LA, Day A, Funnell S, Lovell F, Rob-inson A. Trial of a new acellular pertussis vaccine in healthyadult volunteers. Vaccine 1988;6:29-32.

24 Department of Health. Immunisation against infectiousdisease. London: HMSO,1992:6.

25 Centers for Disease Control and Prevention. FDA approvalof use of a new Haemophilus b conjugate vaccine and acombined diphtheria-tetanus-pertussis and Haemophilus bconjugate vaccine for infants and children. JAMdA 1993;269:2359.

26 Department of Health. Combined Hib/DTP vaccines.CMO's Update 1996-10:1.

27 Ramsay ME, Rao M, Begg NT. Symptoms after acceleratedimmunisation. BMJ 1992;304:1534-6.

on 12 August 2019 by guest. P

rotected by copyright.http://adc.bm

j.com/

Arch D

is Child: first published as 10.1136/adc.75.4.298 on 1 O

ctober 1996. Dow

nloaded from

Vaccines for primary immunisation 303

28 Baraff J, Manclark CR, Cherry JD, Christenson P, MarcySM. Analyses of adverse reactions to diphtheria andtetanus toxoids and pertussis vaccine by vaccine lot, endo-toxin content, pertussis vaccine potency and percentage ofmouse weight gain. Pediatr Infect Dis J 1989;8:502-7.

29 Fritzell B, Plotkin S. Efficacy and safety of a Haemophilusinfluenzae type b capsular polysaccharide-tetanus proteinconjugate vaccine. J Pediatr 1992;121:355-62.

30 Edwards KM, Decker MD, Halsey NA, et al. Differences inantibody response to whole-cell pertussis vaccines. Pediat-

rics 1991;88:1019-23.31 Baker JD, Halperin SA, Edwards K, Miller B, Decker M,

Stephens D. Antibody response to Bordetella pertussisantigens after immunization with American and Canadianwhole-cell vaccines. Pediatr 1992;121:523-7.

32 Barington T, Gyhrs A, Kristensen K, Heilmann C. Oppositeeffects of actively and passively acquired immunity to thecarrier on responses of human infants to a Haemophilusinfluenzae type b conjugate vaccine. Infect Immun 1994;62:9-14.

on 12 August 2019 by guest. P

rotected by copyright.http://adc.bm

j.com/

Arch D

is Child: first published as 10.1136/adc.75.4.298 on 1 O

ctober 1996. Dow

nloaded from