Report of the 1966-67 cholera vaccine field trial in rural East Pakistan*

Bull. Org. mond. Sant) .1969, 40, 199-204Bull. Wid Hithi Org. Report of the 1966-67 Cholera Vaccine Field Trial in Rural East Pakistan* 3. The Lack of Effect of Prior Vaccination or Circulating Vibriocidal Antibody on the Severity of Clinical Cholera WILLIAM M. McCORMACK,' A. S. M. MIZANUR RAHMAN, A. K. M. ALAUDDIN CHOWDHURY, WILEY H. MOSLEY2 & ROBERT A. PHILLIPS3 In a controlled study, it has been shown that the prior administration of cholera vaccine had no beneficial effect on the clinical course of cholera as measured by either the condition of the patient on admission to hospital or the subsequent course of the disease. In fact, the disease was, if anything, more severe in those who had received cholera vaccine. Although the presence ofa high vibriocidal titre is associated with protection from cholera, pre-existing antibody had no effect on the clinical course of the disease in those patients who developed cholera. Vaccination against cholera has been shown to be associated with a reduction in the cholera case rate (Oseasohn et al., 1965; Philippines Cholera Com- mittee, 1965; Benenson et al., 1968a). Serum anti- body, as measured by the vibriocidal test, either naturally occurring or induced by vaccine, has also been shown to be associated with a reduction in the cholera case rate (Mosley et al., 1968a, 1968b). However, it has not been established what effect prior cholera vaccination or the presence of pre- * From the Pakistan-SEATO Cholera Research Labora- tory, Dacca, East Pakistan. The Pakistan-SEATO Cholera Research Laboratory is a part of the SEATO Cholera Re- search Program and is supported by the US Department of State, Agency for International Development; the National Institutes of Health, the National Communicable Disease Center, Public Health Service, US Department of Health, Education, and Welfare; and by the Governments of Pakistan and other SEATO nations. The NIH Cholera Advisory Committee co-ordinates the research programme. These studies were supported in part by Research Agreement No. 196802 between the National Institutes of Health, Bethesda, Md., USA, and the Pakistan-SEATO Cholera Research Laboratory, Dacca, East Pakistan. 1 Formerly Epidemic Intelligence Service Officer, National Communicable Disease Center, Public Health Service, US Department of Health, Education, and Welfare, assigned to Epidemiology Section, Pakistan-SEATO Cholera Research Laboratory. Present Address: Department of Medicine, Massachusetts General Hospital, Boston, Mass., USA. 2Chief, Epidemiology Section, Pakistan-SEATO Cholera Research Laboratory. Director, Pakistan-SEATO Cholera Research Labora- tory. existing antibodies has on the clinical course of cholera in those patients who do develop the disease. Earlier studies to measure the effect of prior cholera vaccination on the cholera case fatality rate provided conflicting results (Pollitzer, 1959). The introduction of effective cholera treatment, based on the replacement of fluid and electrolyte losses (Watten et al., 1959) has reduced the cholera case fatality rate in experienced hands to less than 1% (Lindenbaum et al., 1966). Thus, mortality can no longer be considered to be a reasonable measure of the effect of any factor on the clinical course of cholera. Objective measurements of the severity of clinical cholera can be made. The extent of de- hydration at the time the patient is first seen can be estimated by a determination of serum-protein. The clinical course of the disease after the initiation of treatment can be gauged by the amount of intra- venous fluid required and the duration of diarrhoea. Using these criteria in this study, we have shown that neither prior cholera vaccination nor a pre-existent antibody titre is associated with amelioration of the clinical course of cholera. METHOD The 1966-67 Pakistan-SEATO Cholera Research Laboratory (PSCRL) vaccine field trial has been described in detail by Mosley et al. (1969b). Briefly, 2288 -199- W. M. MCCORMACK AND OTHERS about 40 000 children between the ages of 3 months and 14 years were included. There were 3 vaccine groups: the first group received 2 doses of tetanus and diphtheria toxoids-for adult use(control group); the second group received 1 dose of whole-cell cholera vaccine and 1 dose of tetanus and diphtheria toxoids (1-dose group); the third group was twice as large as the others and received 2 doses of the cholera vaccine (2-dose group). All doses were 0.5 ml. The 2 doses were given at an interval of about 30 days. Only those children who received both assigned doses of cholera vaccine or the control preparation were considered to be participants in the vaccine trial. Neither the field staff nor the hospital staff was aware of the vaccine status of the trial participants. Active surveillance for cholera was carried out in the villages of the vaccine trial area, with field workers visiting each home daily. Rectal swabs for culture were obtained from all individuals under 15 years of age with diarrhoea severe enough to result in a limitation of normal activity. Patients requiring hospitalization were taken to the PSCRL Matlab Bazar Hospital where therapy was administered by a full-time physician (one of the co-authors-A.S.M.M.R.). The standard PSCRL method of cholera treatment, as described by Gordon et al. (1964) was used. A single solution (" 5-4-1 "), containing 5 g of sodium chloride, 4 g of sodium bicarbonate and 1 g of potassium chloride per litre, was administered intravenously, both for the correction of initial dehydration, as estimated clinically, and for the replacement of subsequent fluid losses. Patients were weighed at the time of admission to the hospital and daily thereafter. All patients were treated on " cholera cots " which permitted the collection of liquid stool into calibrated plastic containers. Fluid intake and output were recorded at 8-hourly intervals. The end of diarrhoea was defined as the end of the last 8-hour nursing period during which liquid stool was passed. Rectal swabs for culture were obtained daily from hospitalized patients. Participants in the vac- cine trial were not discharged from hospital until all cultures were negative for Vibrio cholerae for 3 consecutive days. Bacteriological techniques are discussed in detail by Mosley et al. (1969b). A sample of venous blood was obtained on admission to the hospital before initial rehydration. Serum proteins were estimated by refractometry in a Bausch and Lomb serum-protein meter. Serum TABLE 1 PARTICIPANTS WITH DOCUMENTED CHOLERA IN THE PSCRL 1966-67 CHOLERA VACCINE FIELD TRIAL Vaccine group Total Controll 1-dose 2-dose Total cases 35 19 25 79 Field cases 7 4 6 17 Hospital cases 28 15 19 62 Percentage hospitalized 80 78.9 76 78.5 vibriocidal antibody titres were estimated by the microtechnique developed by Benenson et al. (1968b). Antibiotics were not routinely administered. All patients responded well to the intravenous adminis- tration of fluid and electrolytes. In 4 cases where antibiotics were given, either inadvertently or for the treatment of unrelated illnesses, the patients have been excluded from the analysis because of the known modifying effect of antibiotics on the clinical course of cholera (Greenough et al., 1964; Carpenter et al., 1964). RESULTS As shown in Table 1, there were 79 individuals with documented cholera among the vaccine trial participants. Altogether, 17 of these infections were documented in persons with diarrhoea in the field; 16 of these field infections were relatively mild and did not require treatment. The other field case was a child in the control group who died during a severe diarrhoeal illness which lasted only a few hours. Post-mortem cultures obtained from clothing were positive for Vibrio cholerae. The remaining TABLE 2 CHARACTERISTICS OF HOSPITALIZED CHOLERA PATIENTS IN THE PSCRL 1966-67 CHOLERA VACCINE FIELD TRIAL Vaccine group Control I1-dose 2-dose Number of patients 27 12 19 Age-group (years) 0-4 16 10 11 5-9 7 2 7 10-14 4 0 1 Males 14 7 11 Females [ 13 5 8 200 THE 1966-67 CHOLERA VACCINE FIELD TRIAL IN RURAL EAST PAKISTAN. 3 TABLE 3 CHARACTERISTICS REFLECTING SEVERITY OF ILLNESS PRIOR TO HOSPITALIZATION FOR CHOLERA PATIENTS IN THE PSCRL 1966-67 CHOLERA VACCINE FIELD TRIAL Vaccine group Control 1-dose [ 2-dose Number of patients 27 12 19 Mean time between onset of symptoms and hospitalization (hours) 14.9 8.5 16.7 Number of patients without pulse on admission 2 3 7 Mean admission serum-protein (g/100 ml) a 9.2 (26) 10.0 (10) 9.9 (14) Mean percentage gain in weight during hospitalization 7.7 6.3 7.8 a Adequate serum for the determination of serum protein values was not obtained from 8 patients. The numbers in parentheses indicate the number of samples represented by the means. 62 patients were hospitalized. In each vaccine group, about 80% of patients with cholera were hospitalized. Of the 62 hospitalized patients, 4 were excluded from the analysis (3 from the 1-dose group and 1 from the control group) because they received antibiotics. Table 2 lists the age and sex distribution of the 58 hospitalized patients who were included. There were no striking differences. Table 3 lists some clinical characteristics which were indicative of the degree of dehydration at the time of admission to the hospital and reflected the course prior to that time. The mean time between onset of symptoms and admission to the hospital is also listed. It can be seen that more vaccinated individuals than control individuals were without a palpable radial pulse at the time of admission to the hospital (P= <0.01). The degree of dehydration as estimated by the admission plasma protein and the percentage gain in weight after rehydration was similar in all groups. The mean values of several parameters which served as an index of the clinical course of the disease after admission to the hospital are listed in Table 4. All the mean values were higher among those who had received 1 or 2 doses of cholera vaccine, although there were no significant differences in the intravenous fluid requirement, the duration of diarrhoea or the duration of positive bacteriological tests. As a wide range of admission antibody titres was noted among each of the vaccine and control groups TABLE 4 CLINICAL COURSE OF HOSPITALIZED CHOLERA PATIENTS IN THE PSCRL 1966-67 CHOLERA VACCINE FIELD TRIAL Vaccine group Control 1-dose 2-dose Number of patients Mean intravenous fluid requirement (ml/kg of body-weight) Mean duration of diarrhoea (hours) Mean duration of positive bacteriological tests (days) 27 295.9 58.4 12 318.9 72.9 19 450.1 71.2 4.5 5.3 4.8 201 4.5 5.3 4.8 W. M. MCCORMACK AND OTHERS TABLE 5 CLINICAL COURSE OF HOSPITALIZED CHOLERA PATIENTS IN THE PSCRL 1966-67 CHOLERA VACCINE FIELD TRIAL Admission vibriocidal antibody titre (reciprocal) <20 [ 20 40 >40 Number of patients 23 11 11 13 Mean intravenous fluid volume (ml/kg of body-weight) 338.0 352.2 328.0 393.7 Mean duration of diarrhoea (hours) 72.8 61.1 54.4 67.0 Mean duration of positive bacteriolo- gical tests (days) 4.6 4.3 4.2 5.9 (Mosley et al., 1969a), a separate evaluation of the effect of admission antibody titre on the subsequent clinical course of cholera was carried out. The patients were divided into groups on the basis of their admission serum vibriocidal antibody titres, regardless of vaccine status. The criteria used to judge the severity of the clinical course following admission among these groups are compared in Table 5. There was no apparent relationship be- tween vibriocidal antibody titre on admission to hospital and the subsequent clinical course of the disease. DISCUSSION The results of the study indicate quite clearly that there was no beneficial effect from prior cholera vaccination on the clinical course of cholera among hospitalized patients, as measured either by the condition of the patient at the time of admission to the hospital or by the course of the disease during hospitalization. The observation that approximately the same percentage of individuals in each vaccine group required hospitalization is further evidence that the prior administration of vaccine had no beneficial effect on the severity of the disease. A greater proportion of field cases would have been expected in the vaccinated groups if the disease had been modified. Our results are consistent with the observations made by the Philippines Cholera Committee (1966) during their controlled cholera vaccine field trial. They found no difference between the 4 vaccinated groups and the control group in the duration of diarrhoea. Measurements of serum proteins or of intravenous fluid requirements were not available. They concluded that prior vaccination does not alter the course of cholera once the disease has been acquired. As discussed in detail by Mosley et al. (1969a), a rise in titre of vibriocidal antibody is associated with a fall in the clinical cholera case rate. There is, as yet, no evidence to indicate whether or not the vibriocidal antibody is the protective antibody, but it does appear to be a measure of protection. However, among those who developed clinical cholera, we have not been able to show that the presence of a high antibody titre, as measured by the vibriocidal test, had a modifying effect on the course of the illness. Sack et al. (1966) obtained similar results during a study of adult cholera patients in Calcutta. Cholera would then appear to be a disease for which partial immunity fails to influence clinical severity favour- ably. An unexpected observation was that the course of cholera may have been more severe in those patients who had received cholera vaccine. A significantly greater proportion of vaccinated patients were without a palpable radial pulse at the time of hospital admission. In addition, the mean values for all the parameters used to evaluate the clinical course of the disease after hospital admission were greater in the vaccinated groups. The reasons for this are not readily apparent. Perhaps those individuals who, because of exposure to a large inoculum, ordinarily would have had a severe clinical attack, were not protected by vaccine-induced immunity. Volunteer studies in typhoid fever by Hornick & Woodward (1966) offer an interesting parallel. They found that typhoid vaccine was protective only against smaller 202 THE 1966-67 CHOLERA VACCINE FIELD TRIAL IN RURAL EAST PAKISTAN. 3 203 challenge doses. Vaccinated and non-vaccinated subjects developed typical typhoid fever with equal frequency when challenged with larger inocula. If we assume that the clinical severity of cholera in- creases with the size of the inoculum, and if cholera vaccine protects against challenge with small inocula, but not against challenge with large inocula, then we would expect that there would be proportionately more severe cases among those who had received vaccine. ACKNOWLEDGEMENTS Dr Alexander D. Langmuir and Dr Eugene J. Gan- garosa, Epidemiology Program, National Communicable Disease Center, Atlanta, Ga., USA, kindly reviewed the manuscript and offered many helpful suggestions. We acknowledge the invaluable assistance of Mr M. Ibriham, Ward Master, PSCRL Matlab Hospital, and his nursing staff. Mr Sayeedur Rahaman, Statistics Unit, Epidemio- logy Section, PSCRL, provided assistance in processing the data. The vibriocidal titrations were carried out under the supervision of Dr Ansaruddin Ahmed. RESUME RAPPORT SUR L'ESSAI PRATIQUE D'UN VACCIN ANTICHOLERIQUE DANS UNE REGION RURALE DU PAKISTAN ORIENTAL (1966-1967): 3. ABSENCE D'INFLUENCE D'UNE VACCINATION ANTERIEURE OU DE LA PREEXISTENCE D'ANTICORPS VIBRIOCIDES CIRCULANTS SUR LA GRAVITt DE L'EVOLUTION CLINIQUE DU CHOLtRA Dans le cadre de l'essai de vaccin anticholerique dont d'autres aspects sont decrits dans les deux articles prece- dents, on a recherch6 si la vaccination ou l'existence, prealablement it 'infection, d'anticorps vibriocides modi- fiaient les caracteristiques cliniques du cholera. Les observations ont port6 sur 58 malades atteints de cholera confirm6: 12 d'entre eux avaient recu une dose, et 19 deux doses de vaccin antichol6rique; 27 avaient recu une preparation t6moin (anatoxines tetanique et diphterique). Dans chacun de ces groupes, on a procedd a une evaluation clinique de la gravite de la maladie en notant l'intervalle moyen separant l'apparition des sympt6mes et l'hospitalisation, le nombre de patients en etat de choc, la teneur moyenne du serum en proteines et le gain moyen de poids pendant l'hospitalisation. En outre, dans chaque groupe, on a suivi l'evolution de I'affection en se basant sur les criteres suivants: besoins moyens en liquides de remplacement, duree moyenne de la diarrhee et de la positivite des examens bacteriologi- ques. On n'a pas administre systematiquement d'anti- biotiques et tous les malades ont reagi favorablement au traitement rehydratant. L'adoption de ces criteres n'a pas permis de deceler une difference nette d'evolution clinique entre les malades vaccines contre le chol6ra et les malades traites par placebo. De meme, lorsqu'on a group6 les patients, qu'ils aient ete ou non vaccin6s contre le cholera, en fonction des titres d'anticorps seriques qu'ils presentaient au moment de l'admission, on n'a decouvert aucune relation apparente entre la teneur du serum en anti- corps vibriocides et les caracteristiques de l'evolution. II semble qu'une immunite partielle prealable n'exerce aucun effet favorable sur la gravite des manifestations cliniques du cholera. REFERENCES Benenson, A. S., Mosley, W. H., Fahimuddin, M. & Oseasohn, R. 0. (1968a) Bull. WldHith Org., 38,359-372 Benenson, A. S., Saad, A. & Mosley, W. H. (1968b) Bull. Wld Hlth Org., 38, 277-285 Carpenter, C. C. J., Sack, R. B., Mitra, P. P. & Mondal, A. (1964) Bull. Calcutta Sch. trop. Med., 12, 30 Gordon, R. S. et al. (1964) East Pakistan med. J., 8, 10 Greenough, W. B., Gordon, R. S., Rosenberg, I. S., Davies, B. I. & Benenson, A. S. (1964) Lancet, 1, 355 Homick, R. B. & Woodward, T. E. (1966) Trans. Amer. clin. climat. 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