Christophe Rogier

Combating malaria morbidity and mortality by reducing transmission

Jean-François Trape and Christophe Rogier present epidemiological data and an analysis of the relationship between transmission, morbidity and mortality from malaria which suggest that any intervention aiming to reduce transmission will not, on a long-term basis, reduce the burden of malaria in the majority of epidemiological contexts observed in tropical Africa.

Malaria morbidity and pyrethroid resistance after the introduction of insecticide-treated bednets and artemisinin-based combination therapies: a longitudinal study

BACKGROUND Substantial reductions in malaria have been reported in several African countries after distribution of insecticide-treated bednets and the use of artemisinin-based combination therapies (ACTs). Our aim was to assess the effect of these policies on malaria morbidity, mosquito populations, and asymptomatic infections in a west African rural population. METHODS We did a longitudinal study of inhabitants of Dielmo village, Senegal, between January, 2007, and December, 2010. We monitored the inhabitants for fever during this period and we treated malaria attacks with artesunate plus amodiaquine. In July, 2008, we offered longlasting insecticide (deltamethrin)-treated nets (LLINs) to all villagers. We did monthly night collections of mosquitoes during the whole study period, and we assessed asymptomatic carriage from cross-sectional surveys. Our statistical analyses were by negative binomial regression, logistic regression, and binomial or Fisher exact test. FINDINGS There were 464 clinical malaria attacks attributable to Plasmodium falciparum during 17,858 person-months of follow-up. The incidence density of malaria attacks averaged 5·45 (95% CI 4·90-6·05) per 100 person-months between January, 2007, and July, 2008, before the distribution of LLINs. Incidence density decreased to 0·41 (0·29-0·55) between August, 2008, and August, 2010, but increased back to 4·57 (3·54-5·82) between September and December, 2010--ie, 27-30 months after the distribution of LLINs. The rebound of malaria attacks were highest in adults and children aged 10 years or older: 45 (63%) of 71 malaria attacks recorded in 2010 compared with 126 (33%) of 384 in 2007 and 2008 (p<0·0001). 37% of Anopheles gambiae mosquitoes were resistant to deltamethrin in 2010, and the prevalence of the Leu1014Phe kdr resistance mutation increased from 8% in 2007 to 48% in 2010 (p=0·0009). INTERPRETATION Increasing pyrethroid resistance of A gambiae and increasing susceptibility of older children and adults, probably due to decreasing immunity, caused the rebound and age shift of malaria morbidity. Strategies to address the problem of insecticide resistance and to mitigate its effects must be urgently defined and implemented. FUNDING Institut de Recherche pour le Développement and the Pasteur Institute of Dakar.

Long-term clinical protection from falciparum malaria is strongly associated with IgG3 antibodies to merozoite surface protein 3.

Background Surrogate markers of protective immunity to malaria in humans are needed to rationalize malaria vaccine discovery and development. In an effort to identify such markers, and thereby provide a clue to the complex equation malaria vaccine development is facing, we investigated the relationship between protection acquired through exposure in the field with naturally occurring immune responses (i.e., induced by the parasite) to molecules that are considered as valuable vaccine candidates. Methods and Findings We analyzed, under comparative conditions, the antibody responses of each of six isotypes to five leading malaria vaccine candidates in relation to protection acquired by exposure to natural challenges in 217 of the 247 inhabitants of the African village of Dielmo, Senegal (96 children and 121 older adolescents and adults). The status of susceptibility or resistance to malaria was determined by active case detection performed daily by medical doctors over 6 y from a unique follow-up study of this village. Of the 30 immune responses measured, only one, antibodies of the IgG3 isotype directed to merozoite surface protein 3 (MSP3), was strongly associated with clinical protection against malaria in all age groups, i.e., independently of age. This immunological parameter had a higher statistical significance than the sickle cell trait, the strongest factor of protection known against Plasmodium falciparum. A single determination of antibody was significantly associated with the clinical outcome over six consecutive years in children submitted to massive natural parasite challenges by mosquitoes (over three parasite inoculations per week). Finally, the target epitopes of these antibodies were found to be fully conserved. Conclusions Since anti-MSP3 IgG3 antibodies can naturally develop along with protection against P. falciparum infection in young children, our results provide the encouraging indication that these antibodies should be possible to elicit by vaccination early in life. Since these antibodies have been found to achieve parasite killing under in vitro and in vivo conditions, and since they can be readily elicited by immunisation in naïve volunteers, our immunoepidemiological findings support the further development of MSP3-based vaccine formulations.

Combating malaria in Africa

The spread of antimalarial drug resistance has major consequences for malaria control in tropical Africa. Here, the impact of chloroquine resistance on the burden of malaria is analyzed and its implications for the Roll Back Malaria initiative are examined. Malaria mortality has increased at least twofold during the past two decades. Combination therapy should be available for home treatment of young children. The potential toxicity of most antimalarials will require special surveillance programs. The main contribution to malaria control using methods to reduce the entomological inoculation rate is expected in areas with low or unstable transmission. Classic vector-control methods could potentially eliminate malaria in most urban areas and such programs deserve high priority.

Malaria: even more chronic in nature than previously thought; evidence for subpatent parasitaemia detectable by the polymerase chain reaction

In high endemicity areas, malaria is a chronic disease: examination of blood films reveals that up to half of the population, particularly children, harbour parasites at any one given time. The parasitological status of the remainder was addressed using the polymerase chain reaction, a technique 100 to 1000 times more sensitive than microscopy, on a series of samples from Dielmo, a holoendemic area of Senegal. Two-thirds of the microscopically negative individuals were found to harbour subpatent levels of Plasmodium falciparum, suggesting that more than 90% of the exposed population at any one time, i.e. in a cross-sectional survey, are chronically infected. This also means that the range of parasite loads harboured by humans with various degrees of exposure is remarkably large, probably reflecting a large range of effectiveness of the defence mechanisms against malaria parasites, none of which is fully efficient.

Extensive genetic diversity of Plasmodium falciparum isolates collected from patients with severe malaria in Dakar, Senegal

While some genetic host factors are known to protect against severe Plasmodium falciparum malaria, little is known about parasite virulence factors. We have compared the genetic characteristics of P. falciparum isolates collected from 56 severe malaria patients and from 30 mild malaria patients recruited in Hôpital Principal, Dakar, Senegal. All isolates were typed using polymerase chain reaction amplification of polymorphic genetic loci (MSP-1, MSP-2, HRP1, GLURP, CSP, RESA, and the multigene family Pf60). The complexity of infections was lower in severe than in mild malaria and the parasite genetic diversity in both groups was very large. No specific genetic make-up was associated with severity; there were, however, marked differences in allele frequencies in both groups, with a prevalence up to 60% of MSP-2 alleles specifically observed in the severe malaria isolates. In addition, the presence of MSP-1/RO33 alleles was significantly associated with a higher plasma level of tumour necrosis factor alpha receptor 1 (P < 0.05), a reported indicator of severity in human malaria. These results point to potential differences in the genetic characteristics of parasites inducing severe versus mild pathology.

Incidence of tick-borne relapsing fever in west Africa: longitudinal study

BACKGROUND The ongoing drought in sub-Saharan countries has led to the colonisation of west African Savanna by Ornithodoros sonrai; this tick acts as a vector for Borrelia crocidurae, which causes tick-borne relapsing fever (TBRF). Our aim was to ascertain the incidence of TBRF in west Africa. METHODS From 1990 to 2003, we monitored the incidence of TBRF in Dielmo, Senegal, by daily clinical surveillance and by blood testing of individuals with a fever. From 2002 to 2005, we investigated the presence of O sonrai in 30 villages in Senegal, Mauritania, and Mali, and measured by PCR the prevalence of B crocidurae. FINDINGS The average incidence of TBRF over 14 years was 11 per 100 person-years (range from 4 in 1990 to 25 in 1997). All age-groups presented a high incidence of the disease. In addition to relapses, repeated infections in the same individuals were common, with some affected by up to six distinct infections during the study period. Epidemiological studies indicated that 26 of the 30 studied villages (87%) were colonised by the vector tick O sonrai and that the average B crocidurae infection rate of the vector was 31%. INTERPRETATION The incidence of TBRF at the community level is the highest described in Africa for any bacterial disease. The presence of the vector tick in most villages investigated and its high infection rate suggest that TBRF is a common cause of fever in most rural areas of Senegal, Mauritania, and Mali.

5. Variation of Plasmodium falciparum msp1 block 2 and msp2 allele prevalence and of infection complexity in two neighbouring Senegalese villages with different transmission conditions

To investigate the impact of transmission on the development of immunity to malaria and on parasite diversity, longitudinal surveys have been conducted for several years in Dielmo and Ndiop, 2 neighbouring Senegalese villages with holo- and mesoendemic transmission conditions, respectively. We analysed Plasmodium falciparum msp1 block 2 and msp2 genotypes of isolates collected from 58% of the Dielmo villagers during the same week as those studied recently from Ndiop. Allele frequencies differed in both villages, indicating considerable microgeographical heterogeneity of parasite populations. The complexity of the infections, estimated using individual or combined msp1 and msp2 genotyping, in Dielmo was more than double that in Ndiop and it was age-dependent in Dielmo but not in Ndiop. Thus, this study confirmed the influence of age on the complexity of asymptomatic malaria infections in a holoendemic area. The age distribution of complexity in Dielmo substantiates the interpretation that the number of parasite types per isolate reflects acquired antiparasite immunity. This cross-sectional survey also confirms that the sickle cell trait has no impact on complexity but influences the distribution of P. falciparum genotypes.

Combination of malaria vector control interventions in pyrethroid resistance area in Benin: a cluster randomised controlled trial

BACKGROUND Malaria control efforts and elimination in Africa are being challenged by the development of resistance of parasites to antimalarial drugs and vectors to insecticides. We investigated whether the combination of long-lasting insecticidal mosquito nets (LLINs) with indoor residual spraying (IRS) or carbamate-treated plastic sheeting (CTPS) conferred enhanced protection against malaria and better management of pyrethroid-resistance in vectors than did LLINs alone. METHODS We did a cluster randomised controlled trial in 28 villages in southern Benin, west Africa. Inclusion criteria of the villages were moderate level of pyrethroid resistance in malaria vectors and minimum distance between villages of 2 km. We assessed four malaria vector control interventions: LLIN targeted coverage to pregnant women and children younger than 6 years (TLLIN, reference group), LLIN universal coverage of all sleeping units (ULLIN), TLLIN plus full coverage of carbamate-IRS applied every 8 months (TLLIN+IRS), and ULLIN plus full coverage of CTPS lined up to the upper part of the household walls (ULLIN+CTPS). The interventions were allocated to villages by a block randomisation on the basis of preliminary surveys and children of each village were randomly selected to participate with computer-generated numbers. The primary endpoint was the incidence density rate of Plasmodium falciparum clinical malaria in children younger than 6 years as was analysed by Poisson regression taking into account the effect of age and the sampling design with a generalised estimating equation approach. Clinical and parasitological information were obtained by active case detection of malaria episodes during 12 periods of 6 consecutive days scheduled at six weekly intervals and by cross-sectional surveys of asymptomatic plasmodial infections. Children or study investigators were not masked to study group. This study is registered with Current Controlled Trials, number ISRCTN07404145. FINDINGS Of 58 villages assessed, 28 were randomly assigned to intervention groups. 413-429 children were followed up in each intervention group for 18 months. The clinical incidence density of malaria was not reduced in the children from the ULLIN group (incidence density rate 0·95, 95% CI 0·67-1·36, p=0·79), nor in those from the TLLIN+IRS group (1·32, 0·90-1·93, p=0·15) or from the ULLIN+CTPS group (1·05, 0·75-1·48, p=0·77) compared with the reference group (TLLIN). The same trend was observed with the prevalence and parasite density of asymptomatic infections (non significant regression coefficients). INTERPRETATION No significant benefit for reducing malaria morbidity, infection, and transmission was reported when combining LLIN+IRS or LLIN+CTPS compared with a background of LLIN coverage. These findings are important for national malaria control programmes and should help the design of more cost-effective strategies for malaria control and elimination. FUNDING Ministère Français des Affaires Etrangères et Européennes (FSP project 2006-22), Institut de Recherche pour le Développement, Presidents Malaria Initiative (PMI) of US Governement.

A Worldwide Map of Plasmodium falciparum K13-Propeller Polymorphisms.

BACKGROUND Recent gains in reducing the global burden of malaria are threatened by the emergence of Plasmodium falciparum resistance to artemisinins. The discovery that mutations in portions of a P. falciparum gene encoding kelch (K13)-propeller domains are the major determinant of resistance has provided opportunities for monitoring such resistance on a global scale. METHODS We analyzed the K13-propeller sequence polymorphism in 14,037 samples collected in 59 countries in which malaria is endemic. Most of the samples (84.5%) were obtained from patients who were treated at sentinel sites used for nationwide surveillance of antimalarial resistance. We evaluated the emergence and dissemination of mutations by haplotyping neighboring loci. RESULTS We identified 108 nonsynonymous K13 mutations, which showed marked geographic disparity in their frequency and distribution. In Asia, 36.5% of the K13 mutations were distributed within two areas--one in Cambodia, Vietnam, and Laos and the other in western Thailand, Myanmar, and China--with no overlap. In Africa, we observed a broad array of rare nonsynonymous mutations that were not associated with delayed parasite clearance. The gene-edited Dd2 transgenic line with the A578S mutation, which expresses the most frequently observed African allele, was found to be susceptible to artemisinin in vitro on a ring-stage survival assay. CONCLUSIONS No evidence of artemisinin resistance was found outside Southeast Asia and China, where resistance-associated K13 mutations were confined. The common African A578S allele was not associated with clinical or in vitro resistance to artemisinin, and many African mutations appear to be neutral. (Funded by Institut Pasteur Paris and others.).