Molecular markers of Plasmodium falciparum drug resistance in southern highland Rwanda

In Rwanda, frequent mutations in the pfdhfr and pfdhps genes of Plasmodium falciparum have suggested intense sulfadoxine-pyrimethamine resistance. However, data on pfmdr1 are not available but might be important in the context of the first-line treatment with artemether-lumefantrine. During a survey among 749 children under five years of age in southern highland Rwanda, 104 P. falciparum isolates were obtained. Parasite polymorphisms associated with drug sensitivity were typed including the genes pfdhfr, pfdhps, pfmdr1, and pfcrt. Plasma concentrations of chloroquine and pyrimethamine were measured by ELISA. Treatment with artemether-lumefantrine within the preceding two weeks was stated by 12.5% of the respondents; chloroquine in plasma was detected in 17.6%, pyrimethamine in none. Isolates with pfdhfr triple and pfdhps double/triple mutations occurred in 75% and 93%, respectively; 69% of the isolates comprised pfdhfr/pfdhps quintuple or sextuple mutations associated with high-grade sulfadoxine-pyrimethamine resistance. Pfdhfr L164 was absent. The pfmdr1 pattern revealed more than 50% of the F184 polymorphism and almost 40% of the N86-F184-D1246 allele combination known to be selected in infections reappearing following artemether-lumefantrine treatment. Molecular markers demonstrate intense antifolate drug resistance of P. falciparum in southern Rwanda. The present, first-time data on pfmdr1 alleles from Rwanda reveal a pattern which might reflect a predominance of wild types for some alleles or, alternatively, substantial artemether-lumefantrine pressure on the local parasite population.     

Thioctic acid protects against carrageenan-induced acute inflammation in rats by reduction in oxidative stress, downregulation of COX-2 mRNA and enhancement of IL-10 mRNA

Free radicals are highly reactive species which played an important role in the pathogenesis of acute inflammation. In this study, the protective role of the known antioxidant, thioctic acid, in carrageenan-induced acute inflammation in rats was assessed and was compared to the reference non-steroidal anti-inflammatory drug (NSAID) indomethacin. In addition, the roles of oxidative stress, nitric oxide (NO), inducible cyclooxygenase isoform (COX-2) and interleukin (IL)-10 mRNA expressions in thioctic acid-induced effects were also investigated. Inflammation was induced by injection of 0.1 ml of 1.5% carrageenan into the plantar side of right hind paws of the rats. Thioctic acid (50, 100 or 200 mg/kg), indomethacin (10 mg/kg), DMSO and saline were injected i.p. 2 h before carrageenan injection. The percentage increase in paw weight was calculated. Frozen hind paw were used for estimation of lipid peroxides (MDA), NO, GSH, COX-2 and IL-10 mRNA expression. Formalin fixed hind paw were used for histopathological examination. Thioctic acid (200 mg/kg) reduced both paw edema formation and lymphocytes infiltration more significant than indomethacin itself: Both thioctic acid and indomethacin reduced paw MDA and NO formation. In addition, both agents restored the depleted GSH contents in the paw. Thioctic acid decreased the elevated COX-2 mRNA while indomethacin, failed. Furthermore, thioctic acid increased IL-10 mRNA expression while indomethacin decreased its expression. Thioctic acid exhibited a potent anti-inflammatory effect on carrageenan-induced inflammation compared to the NSAID indomethacin. The mechanisms of thioctic induced protection were proved to be due to reduction of NO, MDA, COX-2 mRNA and induction of GSH, IL-10 mRNA.     

Silymarin prevents adriamycin-induced cardiotoxicity and nephrotoxicity in rats

Adriamycin is a potent anticancer agent, its clinical use is limited for its marked cardiotoxicity and nephrotoxicity. The present study aimed to investigate the possible protective role of the natural antioxidant silymarin on ADR-induced heart and kidney toxicity. Studies were performed on four groups of rats. 1--control group, 2--silymarin group (50 mg/kg), 3--adriamycin group (10 mg/kg), 4--adriamycin+silymarin group. On the third day after ADR injection, plasma was separated for determination of LDH, CPK, cholesterol and total lipids. 30 days after ADR injection, plasma was separated for determination of creatinine and urea levels. Frozen heart specimens (72 h) and frozen kidney specimens (30days) were used for estimation of lipid peroxides and GSH contents. Histopathological examinations of heart and kidney sections were also done. Pretreatment of ADR-treated rats with silymarin resulted in a significant decrease in the plasma CPK, LDH, creatinine and urea. On the other hand silymarin pretreatment did not change ADR-induced hyperlipidemia. Silymarin pretreatment significantly decreased the myocardial MDA contents. In addition, silymarin pretreatment normalized renal tissue contents of MDA and GSH. Histopathological examination of heart and kidney sections revealed that ADR caused only mild myocardial injury in silymarin pretreated rats. Also, silymarin pretreatment inhibited ADR-induced renal tubular damage in rats. These results have suggested that, silymarin ameliorated ADR-induced cardiotoxicity and protected against ADR-induced nephrotoxicity in male albino rats. The mechanisms of silymarin induced protection against ADR-induced toxicities were proved to be due to inhibition of lipid peroxidation and protection against GSH depletion.     

A randomised trial to assess the safety and efficacy of artemether-lumefantrine (Coartem) for the treatment of uncomplicated Plasmodium falciparum malaria in Rwanda

Coartem is a fixed-dose combination of artemether-lumefantrine that, given in six doses, provides effective treatment for children with uncomplicated Plasmodium falciparum infection in areas with highly endemic and multidrug-resistant malaria. In Rwanda since 2001, amodiaquine+sulfadoxine-pyrimethamine (AQ+SP) has been the first-line treatment, but resistance to this combination has rapidly emerged and spread. Coartem was considered as a possible alternative, and a randomised, open-label, clinical trial to test its safety, tolerability and efficacy was carried out in 2004-2005. Five hundred children aged 12-59 months with uncomplicated P. falciparum malaria were randomly allocated to AQ+SP or Coartem. Patients were followed up until day 28 after treatment. Adverse events and clinical and parasitological outcomes were recorded. Adequate clinical and parasitological response (ACPR) was significantly higher in children treated with Coartem than in those treated with AQ+SP: the PCR-adjusted 28-day ACPR was 96.68% for Coartem and 79.35% for AQ+SP. Both treatments rapidly cleared parasitaemia and fever, although parasite clearance was significantly faster in children treated with Coartem. Mean packed cell volume increased in all patients, with no significant differences between treatments. Coartem proved to be more efficacious than AQ+SP, with a good safety and tolerability profile.     

Safety and efficacy of dihydroartemisinin/piperaquine (Artekin) for the treatment of uncomplicated Plasmodium falciparum malaria in Rwandan children

In Rwanda, amodiaquine+sulfadoxine/pyrimethamine (AQ+SP) is the current first-line treatment for malaria, introduced in 2001 as an interim strategy before the future deployment of an artemisinin-based combination treatment (ACT). Dihydroartemisinin/piperaquine (DHA-PQP) is a new co-formulated and well tolerated ACT increasingly used in Southeast Asia where it has proved to be highly effective against Plasmodium falciparum malaria. We tested the efficacy, safety and tolerability of DHA-PQP in children with uncomplicated P. falciparum malaria. A randomised, open trial was carried out in 2003-2004. Seven hundred and sixty-two children aged 12-59 months with uncomplicated P. falciparum malaria were randomly allocated to one of the following treatments: amodiaquine+artesunate; AQ+SP; or DHA-PQP. Patients were followed-up until Day 28 after treatment. Adverse events and clinical and parasitological outcomes were recorded. Children treated with DHA-PQP or AQ+AS had a significantly higher cure rate compared with those treated with amodiaquine+sulfadoxine/pyrimethamine (95.2% and 92.0% vs. 84.7%, respectively). Parasite clearance was significantly faster in children treated with DHA-PQP and AQ+AS compared with those treated with amodiaquine+sulfadoxine/pyrimethamine. The frequency of adverse events was significantly lower in patients treated with DHA-PQP than in those treated with combinations containing amodiaquine. A 3-day treatment with DHA-PQP proved to be efficacious with a good safety and tolerability profile and could be a good candidate for the next first-line treatment.     

Standardised versus actual white cell counts in estimating thick film parasitaemia in African children under five

In patients with malaria, parasitaemia is usually estimated by assuming 8000 white cell counts (WCC) per microlitre of blood. In a sample of 3044 African children under 5 years of age with uncomplicated falciparum malaria, parasitaemia estimated using standardised WCC was compared to parasitaemia calculated based on each child's own WCC. The two methods produced comparable results. However, WCC were >8000 in under-fives with an inverse relationship with age, resulting in the standard approximation method significantly underestimating parasitaemia in the youngest age group and overestimating parasitaemia in the oldest age groups.     

Efficacy of amodiaquine alone and combined with sulfadoxine-pyrimethamine and of sulfadoxine pyrimethamine combined with artesunate

The safety and the efficacy of amodiaquine (AQ) alone, AQ plus sulfadoxine-pyrimethamine (SP) (AQ plus SP), and artesunate (ART) plus SP (ART plus SP), three possible alternatives to chloroquine (CQ), were investigated in 379 Rwandan children 6-59 months old with uncomplicated Plasmodium falciparum malaria who visited one urban/peri-urban health center and two rural health centers. The three treatment regimens were well tolerated and no serious adverse effects were observed. Children treated with AQ plus SP had less clinical failures than those treated with ART plus SP (odds ratio [OR] = 0.25, 95% confidence interval [CI] = 0.06-0.81, P = 0.01) or AQ alone (OR = 0.33, 95% CI = 0.07-1.10, P = 0.08). Even after new infections were excluded, AQ plus SP was still significantly more efficacious than ART plus SP (P = 0.05). At day 14, the mean packed cell volume was significantly higher in the AQ plus SP group compared with the ART plus SP group (P = 0.02) and with the AQ alone group (P = 0.01). In Rwanda, AQ plus SP has been chosen to replace CQ as a first-line treatment. However, this is considered an interim measure and new combinations, possibly co-formulated, should be identified and tested.     

Achieving high coverage in Rwanda's national human papillomavirus vaccination programme

Problem

Virtually all women who have cervical cancer are infected with the human papillomavirus (HPV). Of the 275 000 women who die from cervical cancer every year, 88% live in developing countries. Two vaccines against the HPV have been approved. However, vaccine implementation in low-income countries tends to lag behind implementation in high-income countries by 15 to 20 years.

Approach

In 2011, Rwanda’s Ministry of Health partnered with Merck to offer the Gardasil HPV vaccine to all girls of appropriate age. The Ministry formed a “public–private community partnership” to ensure effective and equitable delivery.

Local setting

Thanks to a strong national focus on health systems strengthening, more than 90% of all Rwandan infants aged 12–23 months receive all basic immunizations recommended by the World Health Organization.

Relevant changes

In 2011, Rwanda’s HPV vaccination programme achieved 93.23% coverage after the first three-dose course of vaccination among girls in grade six. This was made possible through school-based vaccination and community involvement in identifying girls absent from or not enrolled in school. A nationwide sensitization campaign preceded delivery of the first dose.

Lessons learnt

Through a series of innovative partnerships, Rwanda reduced the historical two-decade gap in vaccine introduction between high- and low-income countries to just five years. High coverage rates were achieved due to a delivery strategy that built on Rwanda’s strong vaccination system and human resources framework. Following the GAVI Alliance’s decision to begin financing HPV vaccination, Rwanda’s example should motivate other countries to explore universal HPV vaccine coverage, although implementation must be tailored to the local context.