A simple technique for the detection of anti-malarial drug formulations and their presence in human urine.

A simple, sensitive, specific assay technique for the detection and semi-quantification of chloroquine, amodiaquine, quinine, primaquine, sulfadoxine and pyrimethamine in formulations and in human urine by using thin layer chromatography (TLC) was developed and tested in the laboratory. The method involved developing test samples spotted on TLC chromatogram by diethylamine-toluene-isopropanol (1:4:5 v/v/v) as the eluting solvent. The solvent system diethylamine-toluene-isopropanol (1:4:5 v/v/v) enabled the elution and detection of all the tested antimalarial drugs in solution and those spiked in human urine. Detection limits for chloroquine, amodiaquine, quinine and primaquine were the lowest at 0.00025 mg/ml. Sulfadoxine exhibited a detection limit of 0.0005 mg/ml whereas that of pyrimethamine was 0.001 mg/ml. The results indicate the suitability of this technique in antimalarial drug quality and bioavailability studies. It is envisaged that this technique will adequately address the role of drug absorption and excretion in the chemotherapy of malaria as well as to detect types of antimalarial drugs commonly used in the community.

Malaria chemosuppression during pregnancy. VI. Some epidemiological aspects of malaria in infants.

The possible influence of maternal malaria prophylaxis on infancy malaria was assessed in 241 infants. Mothers of 91 infants (PROG-cohort), 99 infants (CQ-cohort) and 51 infants (CQ+PROG-cohort) had received prophylaxis with daily proguanil, once weekly chloroquine, and the two drug combination respectively. Blood smears of infants were examined for parasitaemia once fortnightly. Parasitaemias were treated with either amodiaquine, Fansidar, or Fansidar-quinine combination. In all cohorts, the incidence of malaria parasitaemias within 3 months of age was high (overall mean = 63%). Chloroquine released from its tissue bound form in the CQ and CQ+PROG-cohorts did not have significant chemosuppressive effects on the parasitaemias. Acknowledging that the CQ-prophylaxis group simulated the hypothetical control group, the cohorts similarity in the pattern of parasitaemias suggested that effective maternal malaria chemoprophylaxis during pregnancy did not significantly influence infancy malaria. A sharp rise in incidence around 3 months was indicative of the waning effect of passive immunity. Sole dependence on sub-optimal active immunity led to another sharp rise in incidence from 9 months onwards. The high incidence of infancy malaria parasitaemias calls for increased vigilance in their early detection and effective treatment. Social-cultural factors within the communities may constrain effective disease management.

Malaria chemosuppression in pregnancy. I. The effect of chemosuppressive drugs on maternal parasitaemia.

A randomized prophylactic drug trial was conducted in a malaria holoendemic area, in the Muheza District of Tanzania. Of 327 pregnant women, 124 received proguanil (PROG), 113 chloroquine (CQ), 90 the proguanil and chloroquine combination (CQ+PROG). Prophylaxis was supervised. Chemosuppressive efficacy was measured through the incidence of malaria breakthrough parasitaemias and clinical episodes. Groups were comparable by age, parity, residential area, and enrollment gestational age. Compliance and drug bio-availability was good. The median breakthrough time of the first parasitaemia episode for primigravidae (PG) and multigravidae (MG) was significantly shorter for the CQ group (PG = 56, MG = 78 days) than in the PROG (PG = 97, MG = 112 days) and the CQ+PROG (PG = 138, MG = 140 days) groups. 56% of the CQ group experienced 2 or more clinical episodes compared to 19% (PROG) and 10% (CQ+PROG). PROG and CQ+PROG did not differ significantly. Parasite densities and in vitro tests suggested that CQ selected for more and highly resistant strain(s). Proguanil is suitable for prophylaxis. However, proguanil resistance should be monitored as well as controlled drug distribution and usage. Malaria control strategies other than chemoprophylaxis should be investigated.

Malaria chemosuppression during pregnancy. IV. Its effects on the newborn's passive malaria immunity.

The effect of malaria prophylaxis during pregnancy on the levels of cord blood anti-sporozoite antibodies was investigated in 203 newborns in Muheza, Tanzania. Mothers of 76 newborns had received prophylaxis with proguanil daily (PROG), 66 chloroquine once weekly (CQ), and 61 got a combination of the two drugs (CQ+PROG). Prophylaxis with PROG or CQ+PROG was more efficacious than with CQ. The mean antibody titres were comparable in all three groups, despite titres being significantly low in mothers of the CQ+PROG group. In 93% of 167 paired maternal-cord sera, maternal titres were higher than cord titres. The correlation between maternal and cord titres was low. Parity, placental malaria, and baby maturity showed little influence on titres. Titres of babies delivered by Caesarean section or whose placenta weighed between 0.75 and 1 kg were comparatively low. The first occurrence of a malaria parasitaemia in infants was independent of the levels of cord titres at birth. The results suggested that chemoprophylaxis as effective as PROG or CQ+PROG in holoendemic areas, insignificantly affects maternal-foetal transfer of anti-sporozoite antibodies, and that levels of these antibodies at birth do not modulate the first occurrence of infancy malaria parasitaemia. Interference with the maternal-foetal transfer of this antibody and possibly other component antibodies of passive immunity should not limit the selection of PROG or CQ+PROG for chemoprophylaxis.

Malaria chemosuppression in pregnancy. III. Its effects on the maternal malaria immunity.

The malaria immune status of pregnant women participating in a malaria prophylaxis study was assessed using their sera reactivity to the R32tet32 antigen. Supervised prophylaxis started early in pregnancy till delivery. Women randomly received either chloroquine once weekly (CQ), or proguanil daily (PROG), or a combination of the two drugs (CQ + PROG). Blood was collected at enrollment, then after 8, 16, and 24 weeks of prophylaxis. Of the 312 women who received prophylaxis for more than 10 consecutive weeks before delivery, anti-sporozoite antibodies were assayed in 232 at enrollment, 258 after 8 weeks, and 254 after 16 weeks. Titres at enrollment were comparable by parity and the residential area. Antibodies in women of the PROG group who were parasitaemic before the assessments decreased with the increasing number of breakthrough and clinical episodes. The converse was true for antibodies in the CQ and CQ + PROG groups. Group differences in the parasite densities would explain this. Parity and placental malaria did not influence titres significantly. Overall, antibodies of the CQ + PROG group decrease significantly with time, possibly due to its long period of aparasitaemia. This suggested interference with the immune responsiveness of the women. PROG, which was equally efficacious, offers better prophylaxis.

Malaria chemosuppression in pregnancy. II. Its effect on maternal haemoglobin levels, placental malaria and birth weight.

The malaria prophylactic effects of chloroquine (CQ), proguanil (PROG), and chloroquine-proguanil combination (CQ+PROG) during pregnancy on maternal haemoglobin levels (Hb), placental malaria, and birth weight were assessed in Muheza, Tanzania. Within 2 months of prophylaxis, severe anaemia in primigravidae (PG) was reduced from 21% (22 cases) to 13% (13 cases). There was no positive effect in multigravidae (MG). Sustained increases in the mean Hb occurred in PG of the PROG and CQ+PROG groups. The mean Hb of PG of the CQ group decreased after an initial increase, possibly due to the selection of more and highly chloroquine-resistant strain(s). The mean birth weight of PG was highest in the CQ+PROG (2.89 kg) and least in the CQ group (2.71 kg). The CQ group had the highest low birth weight rate (LBW). The prevalence of placental malaria was highest in the CQ (28%) and lowest in the PROG group (12%). For all the prophylactic effects, PROG and CQ+PROG did not differ significantly. Thence, the deployment of CQ+PROG for prophylaxis would be unnecessary. Proguanil is a suitable alternative to chloroquine prophylaxis. Due to possible emergence of proguanil resistance, deployment of this drug should incorporate constant monitoring for resistance and the eventual prophylaxis efficacy. The search for other effective malaria control measures should continue.

Malaria chemosuppression in pregnancy. V. Placenta malarial changes among three different prophylaxis groups.

The effect of malaria chemoprophylaxis during pregnancy on placenta malarial changes (PMCs) was investigated in 170 tissue sections. Women of 63 sections received daily proguanil (PROG), 61 once weekly chloroquine (CQ) and 46 the two drug combination (CQ+PROG). All were residents of a malaria hyperendemic area in Muheza District, Tanzania. Supervised prophylaxis started early in pregnancy till delivery. Parasitaemias and clinical episodes were detected early and radically treated. Overall, PMCs were mostly infrequent and light viz: fibrinous deposits (98%), fibrinoid necrosis (60%), leucocytic infiltrations (59%), macrophage containing pigment (16%), and malaria parasites (8%). The type, prevalence, and severity of the PMCs in the three prophylaxis groups were comparable. This was despite the fact that PROG and CQ+PROG were prophylactically more efficacious than CQ and despite the expectation that the prevalence and severity of the PMCs would be high in the CQ group. Prompt diagnosis and effective treatment of parasitaemias in this group contributed to the low prevalence and less severity. It is concluded that effective malaria chemoprophylaxis or prompt diagnosis and effective treatment of malaria parasitaemias have significant impact on the prevalence of PMCs. Due to various operational constraints in most developing countries, chemoprophylaxis remains the only feasible broad option for malaria control in pregnancy.

PIP: The study area was Muheza District, north-eastern Tanzania, where some study participants delivering at Muheza District Hospital were matched with non-participants delivering at the same hospital (controls). Comparison of pathologies in the 3 prophylaxis groups was based on the results of 170 randomly selected placenta tissue sections (63 daily proguanil (PROG), 61 once weekly chloroquine (CQ), and 46 CQ+PROG) from women who received prophylaxis for more than 10 consecutive weeks before delivery. The histological features in decreasing order of frequency were: fibrinous deposits (98%), excessive fibrinoid with a possible damage to the villi (villus atrophy) (60%), leucocytic infiltrations (59%), macrophage containing pigment (16%), and lastly malaria-infected red blood cells (8%). Most placenta malarial changes (PCMs) were not associated with major destruction of placenta structures, and parasites did not adhere on the syncytiotrophoblast. Villi atrophy was found in only a few sections with fibrinous deposits. Overall, there were no significant differences in the frequencies of various components of placenta histology by parity and the maternal malaria prophylaxis regimen during pregnancy (p 0.05). In all groups, pathologies were mostly light. The frequency of classes in the 3 prophylaxis groups did not differ significantly (p 0.05). Overall, 133 (78%) of all placentae had no evidence of active or passed infection, while 24 (14%) had signs of past malaria infection and 13 (8%) had signs of active infection. The results of microscopic examination for malaria parasites in the placenta using the placental blood smear were compared with those obtained from the histological examination of the placenta tissue. Only 154 paired histology-blood smear examinations were possible. Examination of placental blood smears revealed twice as many placentae with malaria parasites as those detected by the histological examination of the placenta tissue (p 0.01). The 2 examinations gave the same results in 115 pairs (7 positive and 118 negatives) and differed in 29 pairs (23 smear positive but tissue negative, while 6 were tissue positive but smear negative).

Malaria in infants whose mothers received chemoprophylaxis: response to amodiaquine therapy.

In October 1988, a project was implemented for assessing the malaria chemoprophylactic efficacy of weekly chloroquine (CQ) and daily proguanil (PROG) during pregnancy in Muheza-Tanzania. Resultant CQ and PROG-cohorts of infants were followed up for prompt diagnosis and treatment of malaria. Infections were primarily treated with 25 mg base amodiaquine/kg over 3 days. By September 1990, 49 and 60 infants from PROG and CQ cohorts respectively had completed one year follow up. Thirty-five (71%) infants of PROG and 44 (73%) for CQ-cohort were infected with malaria before 3 months of age. The one year mean infection episode rates were 7 (PROG-cohort) and 6.6 (CQ-cohort). Amodiaquine cleared 209 (80%) of PROG's total infections and 224 (81%) for CQ-cohort, and significantly reduced the infection load among clearance failures. Clearance failures had high pre-treatment parasite densities whilst post-treatment densities were higher in the CQ-cohort than PROG-cohort. Low malaria immunity and chloroquine's long residence time could explain these differences. We conclude that early infancy malaria is common and should always be suspected, looked for and adequately treated. Amodiaquine is better than chloroquine for malaria primary therapy during infancy and early childhood.

Chloroquine therapy still useful in the management of malaria during pregnancy in Muheza, Tanzania.

In searching for effective malaria chemosuppressives during pregnancy in Muheza District--Tanzania, pregnant women are randomly given either 300 mg base chloroquine once weekly or 200 mg daily proguanil. Breakthroughs presenting with clinical malaria are treated with 25 mg base chloroquine/kg (25 CQ) over three days. Due to loss of malaria immunity during pregnancy and Muheza moderate levels and degrees of chloroquine resistance, the in vivo response to 25 CQ was monitored. Between March and May 1989, 49 women were treated resulting into 32 (65%) parasitological clearances and 17 (35%) failures within 7 days. Two of 17 failures (12%) exhibited RIII response and the remaining 15 (88%) had a favourable clinical response. Only 6 (19%) of 32 cleared patients either recrudesced or got reinfected during the three weeks follow up period. In addition to its safety and affordability, the observed drug efficacy during peak malaria transmission and inspite of prevailing resistance makes 25 CQ an ideal first line drug for the management of malaria during pregnancy.