Malaria suppression with fortnightly doses of pyrimethamine with sulfadoxine in the Gambia

Bull. Org. mond. Sante 1970, 43, 513-520 Bull. Wld Hlth Org. Malaria Suppression with Fortnightly Doses of Pyrimethamine with Sulfadoxine in the Gambia A. B. G. LAING 1 As it had been shown in previous trials that very small doses ofpyrimethamine and sulfadoxine or dapsone were effective in eliminating Plasmodium falciparum in 2 or 3 days, small-scale field trials were carried out on children to investigate the efficacy of this combination of drugs in suppressing malaria. Doses of pyrimethamine 2 mg with sulfadoxine 40 mg, fortnightly, appeared to be completely successful in suppressing seasonal hyperendemic malaria in a group of38 Gam- bian children over a 6-month period. On the other hand, pyrimethamine alone at the recommended dosefailed to suppress fakciparum malaria in some children ofanother group. Parasite resistance to the sulfonamide-pyrimethamine combination was not encountered nor was there any suggestion of toxicity. It is concluded that these drugs in combination could have a place in malaria chemoprophylaxis. Clinical trials among hospital patients in the Gambia have shown that potentiating combinations of pyrimethamine and sulfadoxine 2 or dapsone are effective in eliminating Plasmodium falciparum asexual parasitaemias within 2 or 3 days (Laing, 1970). The actual quantity of drugs required to achieve this was found to be extremely small, doses of pyrimethamine as low as 0.01 mg/kg body-weight, combined with a sulfonamide or a sulfone, being effective. Doses of pyrimethamine of 0.1 mg/kg in combination with sulfadoxine were completely effective even in very heavy infections with parasite counts of 500 000 per mm3. These trials were carried out in semi-immune Gambian patients most of whom were infants over 6 months of age or young children under 4 years of age in whom susceptibility to heavy infection was high and death due to malaria not uncommon. The extraordinary efficacy of treatment with these low dosage combinations suggested that they might also be used for suppression: the dosage of sulfon- amide or sulfone would probably be well below a level producing toxic effects and the possibility that parasites would become resistant has been shown by experiments with non-human plasmodia to be un- 1 Medical Research Council Laboratories, Fajara, Bathurst, Gambia. ' Also known as sulphormethoxine (Fanasil). likely." It was decided, therefore, to carry out a small-scale field trial to compare the effects of fortnightly doses of a combination of pyrimethamine and sulfadoxine with weekly doses of pyrimethamine alone over a 6-month period during and just after the rainy season when malaria was hyperendemic. MATERIALS AND METHODS The subjects of the trial were 84 schoolchildren between the ages of 6 and 10 years attending the nearby government school at Bakau, Cape St. Mary, and 36 infants or young children between the ages of 6 months and 4 years from Bakau town. A pre- liminary blood survey was carried out among the schoolchildren in June 1968. Those found with patent parasitaemias were divided into two groups of 26 each; one group received fortnightly pyrimeth- amine 2 mg+sulfadoxine 40 mg and the other pyrimethamine 25 mg weekly; the remaining 32 children had negative blood films and served as a control group who received no antimalarial regimen. The groups of younger children, 12 in each, were made up from those suffering from acute attacks of falciparum malaria attending as out-patients from "Richards, W. H. G. (1968) Resistance of P. berghei etc. to pyrimethamine plus sulphonamides. Paper presented at the Malaria Seminar, London School of Hygiene and Tropical Medicine, May 1968. 2578 -513- A. B. G. LAING June to early September. One group received pyrimethamine 1 mg + sulfadoxine 20 mg for the treatment of the first attack observed and then fortnightly thereafter, and another group pyri- methamine 12.5 mg initially and weekly thereafter; subjects in a third group received treatment for overt malaria occurring during the trial period. Thick blood films stained by Field's rapid method were used to detect or assess parasitaemias every 2 weeks in those groups given suppressive chemo- therapy; in the school control group, blood surveys were carried out only at the beginning of the trial, after 3 months and after 6 months. Parasites were counted against leucocytes and the densities per mmr were estimated accordingly; a thick blood film was reported as negative if no parasites were seen in 100 microscopic fields (magnification x 600). The Medical Research Council grey-wedge photometer was used to estimate haemoglobin con- centrations from finger-prick blood samples. All antimalarial drugs were administered person- ally by the writer. Sulfadoxine was given in the form of Fanasil suspension from drop bottles containing 5 mg of sulfadoxine per drop and pyrimethamine was given as 25-mg Daraprim tablets or Daraprim elixir containing pyrimethamine 1.25 mg per ml. During the 10-week school vacation from the end of July to the end of September, the subjects were examined and treated in their homes; a few were away from the district for varying periods during this time. For the detection of pyrimethamine in urine, 25-ml samples, preserved by the addition of two drops of toluene to each, were sent to the Wellcome Research Laboratories (Beckenham, England) where extracts were examined by thin-layer chromato- graphy. RESULTS Schoolchildren (6-10 years of age) Details of the blood survey results throughout the trial are given in Table 1. Pyrimethamine 2 mg with sulfadoxine 40 mg, fortnightly. Fortnightly doses of this combination were given to 26 schoolchildren. All the children had symptomless parasitaemias at the time of first treatment (22, P. falciparum; 1, P. malariae; 3, mixed): 2 weeks later, all blood films were negative TABLE I NUMBERS OF SCHOOLCHILDREN WITH PARASITAEMIA (July) Week no. (December) Regimen Parasitaemia 1a 3 5 7 9 11 13 15 17 19 21 23 25 _School vacation Pyrimethamine 2 mg P. fakiparum (asexual) 25 - - - - I with sulfadoxine 40 mg, fortnightly P. falciparum (sexual) 3 7 - 1 - - - 1 P. malariae 4 P. ovale Negative - 19 13 18 19 18 22 25 25 26 25 24 25 No. examined 28 26 13 19 19 19 22 26 25 26 25 24 25 Pyrimethamine P. faciparum (asexual) 24 3 2 2 - 1 2 1 3b -_ - - 25 mg, weekly P. falcparum (sexual) 3 4 2 3 1 - - 1 1b P. malariae 4 2 P. ovale 1 - - - - Negative - 18 20 17 14 20 20 24 22 26 25 25 26 No. examined 26 26 24 21 15 21 22 25 25 26 25 25 26 a The children who were given 25 mg of pyrimethamine received their flrst doses In week no. 2. b Treated with pyrimethamine 2 mg + sulfadoxine 40 mg on this occasion. 514 PYRIMETHAMINE AND SULFADOXINE FOR THE SUPPRESSION OF MALARIA IN GAMBIA TABLE 2 HAEMOGLOBIN VALUES (g/100 ml) BEFORE, DURING AND AT THE END OF THE TRIAL No. At 1st treatment After 3 months After 5-6 months Drug regimen of children observed Range Mean Range Mean Range Mean Children 8-10 years of age: pyrimethamine 2 mg with sulfadoxine 40 mg, fortnightly 26 11.0-13.6 12.3 12.4-15.9 14.0 11.1-14.2 12.5 Children 6-10 years of age: pyrimethamine 25 mg, weekly 26 7.5-13.8 11.9 11.6-15.3 13.4 10.8-14.4 12.0 Children 6-10 years of age: control 32 11.1-13.9 12.5 9.3-14.5 12.7 9.9-14.8 12.3 Children 6 months-A years of age: pyrimethamine I mg with sulfadoxine 20 mg, fortnightly 12 5.0-11.3 8.0 10.1-13.2 11.4 except for 7 with P. falciparum gametocytes. There- after, only 3 children were found with patent parasitaemias throughout the trial period. One had a persisting gametocytaemia into the 7th week; another, who had missed 3 consecutive doses, had P. falciparum trophozoites present in the 11th week; and a third, who had missed 5 consecutive doses, had P. falciparum gametocytes present in the 15th week. White-cell counts were done on each subject monthly for the last 3 months of the trial. All counts were within the normal limits although 3 had relatively low counts of 3750, 4000 and 5000 per mms at one or other examination; these counts increased, however, at subsequent examinations and there was no suggestion of leucopenia. Haemoglobin values (see Table 2) increased within the first 3 months from an average of 12.3 g/100 ml to 14.0 g/100 ml and were 12.5 g/100 ml at the end of the trial, similar values to those of children given weekly pyrimethamine. Enlargement of the spleen was found in 19 children at the start of the trial, in 3 after 3 months and in 2 children at the conclusion of the trial. The average dose of pyrimethamine was 0.1 mg/kg body-weight and of sulfadoxine 2.1 mg/kg. Pyrimethamine, 25 mg weekly. Weekly doses were given to 26 schoolchildren, all of whom had symp- tomless parasitaemias (21, P. falciparum; 2, P. mala- riae; 2, mixed infections of these; and 1, P. ovale), and these children were surveyed fortnightly on the same occasions as those given fortnightly doses of the combination. In 3 children asexual falciparum parasitaemia remained patent in spite of the repeated pyrimethamine dosage and in these 3 cases, weekly instead of fortnightly blood films were taken while parasitaemia was patent; details of these infections are given in Table 3. In one child (D3), P. falciparum trophozoites persisted up to the 6th week and although the next 5 doses were missed, the infection appeared to persist subsequently in spite of a resumption of regular dosage. Another child (D17) missed the 3rd dose but asexual parasitaemia cleared after the 5th dose to reappear about a month later, despite regular weekly dosage. In the third child (D18), asexual parasitaemia persisted until after the 6th dose when it disappeared; but further parasitaemia was appar- ent about 2 months later. The presence of pyri- methamine in the urine of all 3 children was con- firmed from urine samples taken in the 17th week, prior to dosage with the combination of pyri- methamine 2 mg + sulfadoxine 40 mg instead of pyrimethamine 25 mg. This combination of drugs apparently eliminated parasitaemia in all 3, as all blood examinations were negative thereafter. Enlarged spleens were detected in 19 children at the start of the trial, in 6 after 3 months and in 4 at the end of the trial. The average dose of pyri- methamine was 1.2 mg/kg. Control group. Thirty-two children who had negative blood films at the time of the first survey were examined again in the 14th week and again at the end of the trial- period. Of 30 children examined at the interim survey, 12 had positive films (11, 515 A. B. G. LAING CL0 CL0 CL0 CL C a. a.o0 a.o a.oL o o Q 0 Q CL O QOQ OC 008 0 OS 008 d le 10 0 0O 0Q8o Q. Q$o _- le _ 0 0 Q0 0Q 0 0 a.0 a. a. 40 0 0 0 a.0 a.oCD . CMI 0- Q. $ v O 0 a). a aC. .0 QL a 40Q) CD a ° 0 O0 a 0 0 V- c-o V QL Qg Qg0 00La D CD 0. 00 r- 00 _ _ C] a a E 0 x 0 70 E C-, 0 coE 0 O QC O 0 0 - n. u C E 0 DcoQ CL P. falciparum; 1, P. ovale) and at the end of the trial period, 11 out of 29 examined were positive (all P. falciparum, 3 being previously negative). Spleen rates at the time of these surveys were 32%, 42% and 41 %, respectively. Two children had anti- malarial treatment for acute attacks of falciparum malaria. Infants and young children (6 months-4 years ofage) All subjects had falciparum malaria when first seen except for one (PS 11) who had quartan malaria; parasitaemias ranged from 1000/mm3 to 240000/mm3. The effects of the two suppressive regimens administered subsequently for periods varying from 4 to 6 months are shown in Table 4. Pyrimethamine I mg with sulfadoxine 20 mg fort- nightly. Twelve infants or young children were treated for acute attacks of falciparum malaria during the early part of the rainy season with a single dose of pyrimethamine 1 mg + sulfadoxine 20 mg. Thereafter, for 5-6 months, they received, approximately fortnightly, the same dose of both drugs, except for 2 subjects who left the district after 1 and 2 months. No detailed follow-up of the immediate response to the first treatment was made, but in all cases the blood films were negative for parasites at the time of the next dose about 2 weeks later, except for 7 showing P. falciparum gameto- cytes; subsequently, except for a few gametocytes in one blood film at the third examination, no parasites were seen throughout the period of observation. Leucocyte counts done in the last week of the trial were all within normal limits. Haemoglobin values (see Table 2) increased from an average of 8.0 g/ 100 ml to 11.4 g/100 ml over the trial period. The average dose of pyrimethamine was 0.1 mg/kg body- weight and of sulfadoxine 2.1 mg/kg. Pyrimethamine 12.5 mg weekly and fortnightly. Twelve infants or young children were treated for acute attacks of falciparum malaria with pyrimeth- amine 12.5 mg and subsequently given the same dose weekly. After 6-8 doses the interval between dosage was increased to fortnightly because of difficulties in arranging weekly attendance of all children. One child (P3) under study for immunological purposes was treated with pyrimethamine 12.5 mg for an acute attack of falciparum malaria a week before being registered for inclusion in this trial and, when still found to have asexual parasitaemia a week later, was referred to the writer; during the previous 9 months the child had been treated with pyrimethamine on 516 LU -Jm Cs uJ D E UN OR w z i I I- 0 Coen z w UJ >Co ILJn co z 0 zLUa -J U 0 0 Iu U)toI z E 0 I- 0. LU-1 -J xX Co N CI2 N VI' CM c0C,' co - r- 1- co X CJ' 0 0 D Ul) CO 00.D =Co PYRIMETHAMINE AND SULFADOXINE FOR THE SUPPRESSION OF MALARIA IN GAMBIA * L U. U L N co le c r- CO c c C c c I. 0. a. a. I.a. a. a. a. a. I0.m I 0 1 IL I' 1 IIL I I LI l 0 04 c z0 u.U)U)U)U)U)U)U)U)C,)U)U)Cf 21 C ._. - 0 0 I, 0 - - 0 - ._ U) E E Iq I I~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ I I II LL 1111o1 co co 0 c UL U. U ,U U. L U. L Ia1 0 I 0 0 0 0 0 IL~ ~IL IL LL _- N C V U 0 |-r a0 °0 m 0. 0. t. l (L M m tl m 9 5 1 z w LU 0 z 0z co C, z 0 U) I- IL 0: 0 04 04 U 0-4 iNZ C#, 04 04 04 04 0 z 0 0, 0 .24 Q -o CD __7_ -cC - 0 E -4 0 E 0-5 co 0 E 0 E E0 CL '7 E 0 0 X 0 co0 E CI Cn U._ I 0 72 0 +0 jD E co 0> Lo. C IL E _i X~ _ a', 0i4, I~ L 11 A. B. G. LAING 2 occasions, apparently successfully. However, on this occasion the infection persisted with continuing schizogony despite increasing the 6th dose of pyri- methamine to 25 mg. In the 7th week, with asexual parasitaemia still present, a dose of the combination of pyrimethamine 1 mg with sulfadoxine 20 mg was given; this appeared to eliminate the infection as no further asexual parasitaemia was found over the subsequent observation period of 3 months. On the fortnightly regimen, one child developed asexual P. falciparum parasitaemia which, however, dis- appeared on increasing the dose to 25 mg. Six of the 12 subjects in this series were away or otherwise not available for 2 months or more. The average dose of pyrimethamine was 1.1 mg/kg. Control group. This consisted of 12 infants or young children from the same village who had overt falciparum malaria during August and early Septem- ber. They were treated with antimalarial drugs, mostly with single therapeutic doses of pyrimeth- amine, and their families were asked to bring them back should they have fever again. In this way, all but one child, whose parents were uncooperative, were seen again before the end of the year with further attacks of malaria-6 with one, 4 with two and 1 with three further attacks. With the possibility of pyrimethamine resistance developing, recrudes- cence cannot be excluded as a cause of these further attacks but reinfection must certainly have occurred. DISCUSSION Compared with the many recent investigations on the effect of combinations of sulfonamide with pyrimethamine or other folic acid antagonists in the treatment of overt malaria, particularly their effec- tiveness in multiple-drug resistant falciparum malaria (Chin et al., 1966; Bartelloni et al., 1967; Herrero, 1967; Sheehy & Reba, 1967), there have been few reports on the effect of such combinations in chemo- prophylaxis. The original antimalarial trials of sulfadoxine in the United Republic of Tanzania included a 6-week trial of weekly 500-mg doses in schoolchildren with and without pyrimethamine 12.5 mg and another trial of sulfadoxine 250 mg with and without pyrimethamine 12.5 mg for 8 weeks (Laing, 1968a). These trials showed that this com- bination was effective against pyrimethamine- resistant falciparum malaria and a further trial in West Malaysia showed that similar doses given weekly eliminated chloroquine-resistant P. falcipa- rum and were also effective against P. vivax (Laing, 1968b). The only other published report to date is that of the trial of weekly sulfadoxine or dapsone with pyrimethamine in 280 Western Nigerian school- children (Lucas et al., 1969). These regimens were successful in suppressing malaria parasitaemias over a 1-year period without serious side-effects and without the emergence of resistant parasites; the doses of sulfadoxine used were 250 mg and 125 mg in combination with pyrimethamine. In the Gambian trial the dose of sulfadoxine was only 40 mg in combination with pyrimethamine 2 mg in schoolchildren and half these quantities in the younger children. These doses, by comparison extremely small, had been shown to be more than sufficient for the treatment of heavy P. falciparum infections in Gambian indigenes (Laing, 1970). In view of the smallness of the doses, the emergence of resistant parasites was thought possible and blood films were taken every 2 weeks from every subject on suppressive regimens. However, no sign of resistance to sulfadoxine with pyrimethamine was found even with fortnightly doses, complete suppression being achieved over the whole trial period of 6 months. This was not so with weekly pyrimethamine in doses of 25 mg which failed to suppress asexual P. fakci- parum parasitaemias in 3 schoolchildren and in 1 infant; however, these pyrimethamine-resistant parasitaemias were sensitive to combined sulfadoxine and pyrimethamine. In the control group of schoolchildren the parasite rate rose from 0% to 40% in 3 months and was at about the same level at the end of the trial, a low rate for the time of year, when more often than not it would be over 60% for this age-group. However, during this wet season there was barely half the usual rainfall so that transmission was almost certainly not as high as usual; nevertheless, as shown in the younger of the two control groups, the possibility of becoming infected was still high as all but one child who did not co-operate in the trial, after the first treatment, returned with one or more acute attacks of falciparum malaria. The over-all fall in haemoglobin levels in the schoolchildren over the last 3 months of malaria suppression, both in those given the combination and those given pyrimethamine alone, is difficult to explain. The removal of patent asexual parasitaemia with consequent haemolysis would stimulate reticulo- cytosis and red cell regeneration so that an increased haemoglobin level could be expected and this is reflected in the raised levels after 3 months of 518 PYRIMETHAMINE AND SULFADOXINE FOR THE SUPPRESSION OF MALARIA IN GAMBIA suppression. Thereafter, instead of persisting at a high level, a reversion to the same level as that found in the control group appears to have taken place. The reason for such a reversion on the data available is not clear but the finding of the same levels of haemoglobin in both parasitized and non- parasitized groups of children suggests that in the former the rate of red-cell destruction is fully compensated by their rate of replacement. The successful outcome of this trial, together with that of the more comprehensive Nigerian trial, shows that combinations of sulfonamide and pyri- methamine are highly effective in suppressing falciparum malaria in indigenous African popula- tions, more so in fact than pyrimethamine alone. In neither trial did any side-effects occur, although in the Nigerian trial sulfadoxine may have been re- sponsible for transient anaemia associated with glucose-6-phosphate dehydrogenase deficiency in 2 children and leucopenia in 3 children; nor was there any suggestion that the parasites became resistant to the drugs in combination. However, bacteriological examination of stool specimens in the Nigerian trial suggested that sulfonamide-resistant strains of Escherichia coli were detected in the first 3 months of the trial but these did not become more prevalent later. Further investigations on this aspect of the use of sulfonamides in malaria chemo- prophylaxis are indicated but there would appear to be no objection to the inclusion of sulfadoxine in combination with pyrimethamine among the recom- mended agents for the prevention of malaria. ACKNOWLEDGEMENTS I am grateful to Mr Abdoulie K. Bojang for his cheer- ful assistance with both the laboratory routine of this investigation and the maintenance of co-operation among the trial subjects and their families. My thanks are also due to the Wellcome Research Laboratories, particularly to Mr C. R. Jones of the Biophysics and Biochemistry Department, for carrying out the pyrimethamine-urine tests. I wish to thank Dr I. A. McGregor, Director, Medical Research Council Laboratories, the Gambia, for per- mission to publish the results of this trial. RESUMlt TRAITEMENT SUPPRESSIF DU PALUDISME PAR DES DOSES BIMENSUELLES DE PYRIMtTHAMINE ASSOCItE A LA SULFADOXINE, EN GAMBIE Des essais cliniques effectues en Gambie sur des palu- deens semi-immuns hospitalises ont montre que l'admi- nistration de pyrimethamine associee a la sulfadoxine ou au dapsone, qui la potentialisent, permet d'eliminer en 2 ou 3 jours une forte parasitemie a Plasmodium falci- parum. La quantite de medicaments necessaire etait extremement faible: associ6es a un sulfamide ou a une sulfone, des doses de pyrimethamine n'excedant meme pas 0,01 mg/kg de poids corporel venaient a bout d'une parasitemie a formes asexu6es. L'extraordinaire efficacit6 de faibles doses de ces associations de medicaments a conduit a penser qu'elles seraient peut-etre utilisables egalement dans le traitement suppressif. On a donc compar6 l'effet suppressif de doses de 2 mg de pyrimethamine + 40 mg de sulfadoxine, administrees tous les 15 jours pendant 6 mois, it celui de 25 mg de pyrimethamine seule donnes une fois par semaine, pendant le meme laps de temps, chez des enfants gam- biens, ag6s de 6 a 10 ans, atteints de parasitemie asympto- matique a P. falciparum. Avec l'association medicamen- teuse, on a pratiquement obtenu une suppression totale de la parasit6mie apres la premiere dose, alors que chez 3 des enfants traites chaque semaine par la pyrimetha- mine seule persistait une parasitemie it formes asexuees de P. falciparum, qu'une dose de l'association medica- menteuse fit, par la suite, disparaitre. De meme, on a compare l'effet suppressif de doses de 1 mg de pyrimdthamine + 20 mg de sulfadoxine, admi- nistrees tous les 15 jours pendant 5 ou 6 mois, a celui de 12,5 mg de pyrim6thamine seule donnes une fois par semaine, puis tous les 15 jours, chez des enfants gam- biens ages de 6 mois a 3½/2 ans. A nouveau, 1'effet sup- pressif de l'association medicamenteuse a ete total, alors que chez un des enfants traites chaque semaine par la pyrimethamine seule persistait une parasitemie a formes asexu6es de P. falciparum, qui disparut apres l'adminis- tration d'une dose de l'association medicamenteuse. L'examen periodique de groupes temoins a mis en evidence que le niveau de transmission du paludisme etait reste eleve pendant toute la periode des essais. 519 520 A. B. G. LAING REFERENCES Bartelloni, P. J., Sheehy, T. W. & Tigertt, W. D. (1967) J. Amer. med. Ass., 199, 141 Chin, W. et al. (1966) Amer. J. trop. Med. Hyg., 15, 823 Herrero, J. (1967) Rev. Soc. bras. 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