Multi-site comparison of factors influencing progress of African insecticide testing facilities towards an international Quality Management System certification.

BACKGROUND: Insecticidal mosquito vector control products are vital components of malaria control programmes. Test facilities are key in assessing the effectiveness of vector control products against local mosquito populations, in environments where they will be used. Data from these test facilities must be of a high quality to be accepted by regulatory authorities, including the WHO Prequalification Team for vector control products. In 2013-4, seven insecticide testing facilities across sub-Saharan Africa, with technical and financial support from Innovative Vector Control Consortium (IVCC), began development and implementation of quality management system compliant with the principles of Good Laboratory Practice (GLP) to improve data quality and reliability. METHODS AND PRINCIPLE FINDINGS: We conducted semi-structured interviews, emails, and video-call interviews with individuals at five test facilities engaged in the IVCC-supported programme and working towards or having achieved GLP. We used framework analysis to identify and describe factors affeting progress towards GLP. We found that eight factors were instrumental in progress, and that test facilities had varying levels of control over these factors. They had high control over the training programme, project planning, and senior leadership support; medium control over infrastructure development, staff structure, and procurement; and low control over funding the availability and accessibility of relevant expertise. Collaboration with IVCC and other partners was key to overcoming the challenges associated with low and medium control factors. CONCLUSION: For partnership and consortia models of research capacity strengthening, test facilities can use their own internal resources to address identified high-control factors. Project plans should allow additional time for interaction with external agencies to address medium-control factors, and partners with access to expertise and funding should concentrate their efforts on supporting institutions to address low-control factors. In practice, this includes planning for financial sustainability at the outset, and acting to strengthen national and regional training capacity.

Evaluation of ICON Maxx, a long-lasting treatment kit for mosquito nets: experimental hut trials against anopheline mosquitoes in Tanzania.

BACKGROUND: Insecticide-treated nets are the primary method of preventing malaria. To remain effective, the pyrethroid insecticide must withstand multiple washes over the lifetime of the net. ICON(®) Maxx is a 'dip-it-yourself' kit for long-lasting treatment of polyester nets. The twin-sachet kit contains a slow-release capsule suspension of lambda-cyhalothrin plus binding agent. To determine whether ICON Maxx meets the standards required by the World Health Organization Pesticide Evaluation Scheme (WHOPES), the efficacy and wash fastness of ICON Maxx was evaluated against wild, free-flying anopheline mosquitoes. METHODS: ICON Maxx was subjected to bioassay evaluation and experimental hut trial against pyrethroid-susceptible Anopheles gambiae, Anopheles arabiensis and Anopheles funestus. Mosquito mortality, blood feeding inhibition and personal protection were compared between untreated nets, conventional lambda-cyhalothrin treated nets (CTN) washed either four times (cut-off threshold) or 20 times, and ICON Maxx-treated nets either unwashed or washed 20 times. RESULTS: In bioassay, ICON Maxx demonstrated superior wash resistance to the CTN. In the experimental hut trial, ICON Maxx killed 75 % of An. funestus, 71 % of An. gambiae and 47 % of An. arabiensis when unwashed and 58, 66 and 42 %, respectively, when 20 times washed. The CTN killed 52 % of An. funestus, 33 % of An. gambiae and 30 % of An. arabiensis when washed to the cut-off threshold of four washes and 40, 40 and 36 %, respectively, when 20 times washed. Percentage mortality with ICON Maxx 20 times washed was similar (An. funestus) or significantly higher (An. gambiae, An. arabiensis) than with CTN washed to the WHOPES cut-off threshold. Blood-feeding inhibition with ICON Maxx 20 times washed was similar to the CTN washed to cut-off for all three species. Personal protection was significantly higher with ICON Maxx 20 times washed (66-79 %) than with CTN washed to cut-off (48-60 %). CONCLUSIONS: Nets treated with ICON Maxx and washed 20 times met the approval criteria set by WHOPES for Phase II trials in terms of mortality and blood-feeding inhibition. This finding raises the prospect of conventional polyester nets and other materials being made long-lastingly insecticidal through simple dipping in community or home, and thus represents a major advance over conventional pyrethroid treatments.

α-Thalassaemia trait is associated with antibody prevalence against malaria antigens AMA-1 and MSP-1.

A longitudinal study was conducted in a low endemic area in northern Tanzania to examine the influence of the α-thalassaemia trait on malaria incidence and antibody responses to malaria apical membrane antigen-1 (AMA-1) and merozoite surface protein1-19 (MSP-119). Out of 394 children genotyped for α-thalassaemia trait, 4.1% (16 of 394) and 30.7% (121 of 394) were homozygous and heterozygous, respectively. During the 1 year follow-up, four incidents of malaria cases were detected without an evident association with α-thalassaemia. Being heterozygous or homozygous for α-thalassaemia was associated with an increased prevalence of antibodies to AMA-1 [odds ratio (OR): 1.83, 95% confidence interval (CI): 1.07-3.12, p = 0.027] and MSP-1 (OR: 2.04, 95% CI: 1.16-3.60, p = 0.013) after adjustment for age and reported bednet use. The observed association between α-thalassaemia and malaria antibody responses may reflect longer-term differences in antigen exposure or differences in antibody acquisition upon exposure in this low endemic setting.

Trends in chloroquine resistance marker, Pfcrt-K76T mutation ten years after chloroquine withdrawal in Tanzania.

BACKGROUND: Plasmodium falciparum resistance to anti-malarial drugs remains a major obstacle to the control of malaria. In 2001 Tanzania replaced chloroquine (CQ) with sulphadoxine-pyrimethamine (SP) as first-line drug, which in turn was replaced by artemisinin combination therapy in 2006. SP has however, continued to be used in intermittent preventive treatment of malaria in pregnancy (IPTp) despite reports of high levels of resistance to SP due to the lack of alternatives to SP for IPTp. Recent reports have indicated recovery of CQ-susceptibility in Malawi, Kenya, Mozambique, and Tanzania based on the prevalence of wild types at codon 76 of the Pfcrt gene in indigenous P. falciparum populations. The current prevalence of this Pfcrt-76 CQ resistance marker from six regions of Tanzania mainland is hereby reported. METHODS: DNA extracted from filter-paper dried blood spots and rapid diagnostics kit strips collected from finger-prick blood were used to genotype the Pfcrt-76 resistance marker using PCR-RFLP. Data from previously published studies were used to generate CQ susceptibility recovery trends using logistic regression model. RESULTS: Seven hundred and forty one (741) samples were genotyped. The current frequency of the CQ-susceptible Pfcrt-K76 was above 92% and did not differ between regions in Tanzania (χ(2) = 2.37; p = 0.795). The K76 allelic prevalence was between 85.7 and 93% in regions (χ(2) = 7.88, p = 0.163). The CQ resistance recovery trends showed regional variability that may be caused by differences in malaria transmission intensity, but overall the trends converge as the susceptibility levels in all regions approach >90%. CONCLUSIONS: CQ withdrawal in Tanzania has resulted into >90% recovery of susceptibility in ten years of withdrawal. These findings are in support of the search for CQ-based combination drugs as a possible future alternative to SP for IPTp in places where full recovery of CQ-susceptibility will be evident.

A cluster-randomized trial of mass drug administration with a gametocytocidal drug combination to interrupt malaria transmission in a low endemic area in Tanzania.

BACKGROUND: Effective mass drug administration (MDA) with anti-malarial drugs can clear the human infectious reservoir for malaria and thereby interrupt malaria transmission. The likelihood of success of MDA depends on the intensity and seasonality of malaria transmission, the efficacy of the intervention in rapidly clearing all malaria parasite stages and the degree to which symptomatic and asymptomatic parasite carriers participate in the intervention. The impact of MDA with the gametocytocidal drug combination sulphadoxine-pyrimethamine (SP) plus artesunate (AS) plus primaquine (PQ, single dose 0.75 mg/kg) on malaria transmission was determined in an area of very low and seasonal malaria transmission in northern Tanzania. METHODS: In a cluster-randomized trial in four villages in Lower Moshi, Tanzania, eight clusters (1,110 individuals; cluster size 47- 209) were randomized to observed treatment with SP+AS+PQ and eight clusters (2,347 individuals, cluster size 55- 737) to treatment with placebo over three days. Intervention and control clusters were 1 km apart; households that were located between clusters were treated as buffer zones where all individuals received SP+AS+PQ but were not selected for the evaluation. Passive case detection was done for the entire cohort and active case detection in 149 children aged 1-10 year from the intervention arm and 143 from the control arm. Four cross-sectional surveys assessed parasite carriage by microscopy and molecular methods during a five-month follow-up period. RESULTS: The coverage rate in the intervention arm was 93.0% (1,117/1,201). Parasite prevalence by molecular detection methods was 2.2-2.7% prior to the intervention and undetectable during follow-up in both the control and intervention clusters. None of the slides collected during cross-sectional surveys had microscopically detectable parasite densities. Three clinical malaria episodes occurred in the intervention (n = 1) and control clusters (n = 2). CONCLUSIONS: This study illustrates the possibility to achieve high coverage with a three-day intervention but also the difficulty in defining suitable outcome measures to evaluate interventions in areas of very low malaria transmission intensity. The decline in transmission intensity prior to the intervention made it impossible to assess the impact of MDA in the chosen study setting. TRIAL REGISTRATION: ClinicalTrials.gov: NCT00509015.

Policy environment and male circumcision for HIV prevention: findings from a situation analysis study in Tanzania.

BACKGROUND: Male circumcision (MC) has been shown to be effective against heterosexual acquisition of HIV infection and is being scaled up as an additional strategy against HIV in several countries of Africa. However, the policy environment (whether to formulate new specific policy on MC or adapts the existing ones); and the role of various stakeholders in the MC scale up process in Tanzania was unclear. We conducted this study as part of a situation analysis to understand the attitudes of policy makers and other key community and health authority decision makers towards MC, policy and regulatory environment, and the readiness of a health system to accommodate scaling up of MC services. METHODS: We conducted 36 key informants' interviews with a broad range of informants including civil servants, religious leaders, cultural and traditional gatekeepers and other potential informants. Study informants were selected at the national level, regional, district and community levels to represent both traditionally circumcising and non-circumcising communities. RESULTS: Study informants had positive attitudes and strong beliefs towards MC. Key informants in traditionally non-circumcising districts were willing to take their sons for medically performed MC. Religious leaders and traditional gatekeepers supported MC as it has been enshrined in their holy scripts and traditional customs respectively. Civil servants highlighted the need for existence of enabling policy and regulatory environment in the form of laws, regulations and guidelines that will ensure voluntary accessibility, acceptability, quality and safety for those in need of MC services. Majority of informants urged the government to make improvements in the health system at all levels to ensure availability of adequate trained personnel, infrastructure, equipment, and supplies for MC scale up, and insisted on the involvement of different MC stakeholders as key components in effective roll out of medically performed MC programme in the country. CONCLUSIONS: Findings from the situation analysis in Tanzania have shown that despite the absence of a specific policy on MC, basic elements of enabling policy environment at national, regional, district and community levels are in place for the implementation of MC scale up programme.

Acceptability of medical male circumcision in the traditionally circumcising communities in Northern Tanzania.

BACKGROUND: Data from traditionally circumcising communities show that non-circumcised males and those circumcised in the medical settings are stigmatised. This is because traditional circumcision embodies local notions of bravery as anaesthetics are not used. This study was conducted to assess the acceptability of safe medical circumcision before the onset of sexual activity for HIV infection risk reduction in a traditionally circumcising community in Tanzania. METHODS: A cross-sectional study was conducted among males and females aged 18-44 years in traditionally circumcising communities of Tarime District in Mara Region, North-eastern Tanzania. A face-to-face questionnaire was administered to females to collect information on the attitudes of women towards circumcision and the preferred age for circumcision. A similar questionnaire was administered to males to collect information on socio-demographic, preferred age for circumcision, factors influencing circumcision, client satisfaction, complications and beliefs surrounding the practice. RESULTS: Results were available for 170 males and 189 females. Of the males, 168 (98.8%) were circumcised and 61 (36.3%) of those circumcised had the procedure done in the medical setting. Of those interviewed, 165 (97.1%) males and 179 (94.7%) females supported medical male circumcision for their sons. Of these, 107 (64.8%) males and 130 (72.6%) females preferred prepubertal medical male circumcision (12 years or less). Preference for prepubertal circumcision was significantly associated with non-Kurya ethnic group, circumcision in the medical setting and residence in urban areas for males in the adjusted analysis. For females, preference for prepubertal circumcision was significantly associated non-Kurya ethnic group and being born in urban areas in the adjusted analysis. CONCLUSIONS: There is a shift of preference from traditional male circumcision to medical male circumcision in this traditionally circumcising population. However, this preference has not changed the circumcision practices in the communities because of the community social pressure. Male circumcision national program should take advantage of this preference of medical male circumcision by introducing safe and affordable circumcision services and mobilising communities in a culturally sensitive manner to take up circumcision services.

Traditional male circumcision practices among the Kurya of North-eastern Tanzania and implications for national programmes.

The World Health Organisation and the Joint United Nations Programme on AIDS recommend male circumcision (MC) as an additional intervention against HIV infection. Various sub-Saharan African countries are at different stages of rolling out MC programmes. Despite initial fears, studies conducted among traditionally non-circumcising communities in Africa have shown that MC is widely accepted as a biomedical intervention. However, little is known on how traditionally circumcising communities where MC carries considerable social meaning and significance would respond to such programmes. This study was conducted among a traditionally circumcising community in Tarime district in Tanzania as part of a national situation analysis prior to initiating a national MC programme. It employed key informant interviews and focus group discussions for data collection. Results show that the Kurya ethnic group practice MC as a rite of passage from childhood to adulthood. Each clan organises its own circumcision ceremony, which takes place every even numbered years. Clan leaders and traditional circumcisers are central to its organisation. Among the Kurya, there is high regard for traditional MC as it is perceived as upholding cultural practice and identity. It also embodies notions of bravery since anaesthetics are not used. On the other hand, medical MC is not viewed as prestigious since anaesthetics are used to suppress pain. Social pressure for traditional MC is applied through ridiculing of those uncircumcised or circumcised at health facilities. In general, there are positive attitudes towards MC as it is perceived as enhancing personal hygiene and having a protective effect against sexually transmitted infections. For the success of nation-wide MC programmes, there is need to develop programmes that incorporate both clinical and sociocultural interests.

The dynamics of naturally acquired immune responses to Plasmodium falciparum sexual stage antigens Pfs230 & Pfs48/45 in a low endemic area in Tanzania.

BACKGROUND: Naturally acquired immune responses against sexual stages of P. falciparum can reduce the transmission of malaria from humans to mosquitoes. These antigens are candidate transmission-blocking vaccines but little is known about the acquisition of sexual stage immunity after exposure to gametocytes, or their longevity and functionality. We conducted a longitudinal study on functional sexual stage immune responses. METHODOLOGY/PRINCIPAL FINDINGS: Parasitaemic individuals (n = 116) were recruited at a health centre in Lower Moshi, Tanzania. Patients presented with gametocytes (n = 16), developed circulating gametocytes by day 7 (n = 69) or between day 7 and 14 (n = 10) after treatment or did not develop gametocytes (n = 21). Serum samples were collected on the first day of gametocytaemia and 28 and 84 days post-enrolment (or d7, 28, 84 after enrolment from gametocyte-negative individuals). Antibody responses to sexual stage antigens Pfs230 and Pfs48/45 were detected in 20.7% (72/348) and 15.2% (53/348) of the samples, respectively, and were less prevalent than antibodies against asexual stage antigens MSP-1(19) (48.1%; 137/285) and AMA-1 (52.4%; 129/246)(p<0.001). The prevalence of anti-Pfs230 (p = 0.026) and anti-Pfs48/45 antibodies (p = 0.017) increased with longer duration of gametocyte exposure and had an estimated half-life of approximately 3 months. Membrane feeding experiments demonstrated a strong association between the prevalence and concentration of Pfs230 and Pfs48/45 antibodies and transmission reducing activity (TRA, p<0.01). CONCLUSIONS/SIGNIFICANCE: In a longitudinal study, anti-Pfs230 and Pfs48/45 antibodies developed rapidly after exposure to gametocytes and were strongly associated with transmission-reducing activity. Our data indicate that the extent of antigen exposure is important in eliciting functional transmission-reducing immune responses.

Identification of hot spots of malaria transmission for targeted malaria control.

BACKGROUND: Variation in the risk of malaria within populations is a frequently described but poorly understood phenomenon. This heterogeneity creates opportunities for targeted interventions but only if hot spots of malaria transmission can be easily identified. METHODS: We determined spatial patterns in malaria transmission in a district in northeastern Tanzania, using malaria incidence data from a cohort study involving infants and household-level mosquito sampling data. The parasite prevalence rates and age-specific seroconversion rates (SCRs) of antibodies against Plasmodium falciparum antigens were determined in samples obtained from people attending health care facilities. RESULTS: Five clusters of higher malaria incidence were detected and interpreted as hot spots of transmission. These hot spots partially overlapped with clusters of higher mosquito exposure but could not be satisfactorily predicted by a probability model based on environmental factors. Small-scale local variation in malaria exposure was detected by parasite prevalence rates and SCR estimates for samples of health care facility attendees. SCR estimates were strongly associated with local malaria incidence rates and predicted hot spots of malaria transmission with 95% sensitivity and 85% specificity. CONCLUSIONS: Serological markers were able to detect spatial variation in malaria transmission at the microepidemiological level, and they have the potential to form an effective method for spatial targeting of malaria control efforts.

Duration of protection against clinical malaria provided by three regimens of intermittent preventive treatment in Tanzanian infants.

BACKGROUND: Intermittent preventive treatment in infants (IPTi) is a new malaria control tool. However, it is uncertain whether IPTi works mainly through chemoprophylaxis or treatment of existing infections. Understanding the mechanism is essential for development of replacements for sulfadoxine-pyrimethamine (SP) where it is no longer effective. This study investigated how protection against malaria given by SP, chlorproguanil-dapsone (CD) and mefloquine (MQ), varied with time since administration of IPTi. METHODS AND FINDINGS: A secondary analysis of data from a randomised, placebo-controlled trial in an area of high antifolate resistance in Tanzania was conducted. IPTi using SP, CD, MQ or placebo was given to 1280 infants at 2, 3 and 9 months of age. Poisson regression with random effects to adjust for potential clustering of malaria episodes within children was used to calculate incidence rate ratios for clinical malaria in defined time strata following IPTi. The short-acting antimalarial CD gave no protection against clinical malaria, whereas long-acting MQ gave two months of substantial protection (protective efficacy (PE) 73.1% (95% CI: 23.9, 90.5) and 73.3% (95% CI: 0, 92.9) in the first and second month respectively). SP gave some protection in the first month after treatment (PE 64.5% (95% CI: 10.6, 85.9)) although it did not reduce the incidence of malaria up to 12 months of age. There was no evidence of either long-term protection or increased risk of malaria for any of the regimens. CONCLUSION: Post-treatment chemoprophylaxis appears to be the main mechanism by which IPTi protects children against malaria. Long-acting antimalarials are therefore likely to be the most effective drugs for IPTi, but as monotherapies could be vulnerable to development of drug resistance. Due to concerns about tolerability, the mefloquine formulation used in this study is not suitable for IPTi. Further investigation of combinations of long-acting antimalarials for IPTi is needed. TRIAL REGISTRATION: Clinicaltrials.gov NCT00158574.

Protective efficacy and safety of three antimalarial regimens for intermittent preventive treatment for malaria in infants: a randomised, double-blind, placebo-controlled trial.

BACKGROUND: Administration of sulfadoxine-pyrimethamine at times of vaccination-intermittent preventive treatment in infants (IPTi)-is a promising strategy to prevent malaria. However, rising resistance to this combination is a concern. We investigated a shortacting and longacting antimalarial drug as alternative regimens for IPTi. METHODS: We undertook a double-blind, placebo-controlled trial of IPTi in an area of high resistance to sulfadoxine-pyrimethamine at sites of moderate (n=1280 infants enrolled) and low (n=1139) intensity of malaria transmission in Tanzania. Infants aged 8-16 weeks were randomly assigned in blocks of 16 to sulfadoxine (250 mg) plus pyrimethamine (12.5 mg; n=319 in moderate-transmission and 283 in low-transmission sites), chlorproguanil (15 mg) plus dapsone (18.75 mg; n=317 and 285), mefloquine (125 mg; n=320 and 284), or placebo (n=320 and 284), given at the second and third immunisations for diphtheria, pertussis, and tetanus, and for measles. Research team and child were masked to treatment. Recruitment was stopped early at the low-transmission site because of low malaria incidence. The primary endpoint was protective efficacy against all episodes of clinical malaria at 2-11 months of age. Analysis was by intention to treat. This study is registered with ClinicalTrials.gov, number NCT00158574. FINDINGS: All randomly assigned infants were analysed. At the moderate-transmission site, mefloquine had a protective efficacy of 38.1% (95% CI 11.8-56.5, p=0.008) against clinical malaria in infants aged 2-11 months, but neither sulfadoxine-pyrimethamine (-6.7%, -45.9 to 22.0) nor chlorproguanil-dapsone (10.8%, -24.6 to 36.1) had a protective effect. No regimen had any protective efficacy against anaemia or hospital admission. Mefloquine caused vomiting in 141 of 1731 (8%) doses given on day 1 (odds ratio vs placebo 5.50, 95% CI 3.56-8.46). More infants died in the chlorproguanil-dapsone and mefloquine groups (18 and 15, respectively) than in the sulfadoxine-pyrimethamine or placebo groups (eight deaths per group; p=0.05 for difference between chlorproguanil-dapsone and placebo). INTERPRETATION: IPTi with a longacting, efficacious drug such as mefloquine can reduce episodes of malaria in infants in a moderate-transmission setting. IPTi with sulfadoxine-pyrimethamine has no benefit in areas of very high resistance to this combination. The appropriateness of IPTi should be measured by the expected incidence of malaria and the efficacy, tolerability, and safety of the drug. FUNDING: IPTi Consortium and the Gates Malaria Partnership.

Common genotypic polymorphisms in glutathione S-transferases in mild and severe falciparum malaria in Tanzanian children.

Malaria infection induces oxidative stress in the host cells. Antioxidant enzymes such as glutathione S-transferases (GSTs) are responsible for fighting reactive oxygen species and reduction of oxidative stress. Common GST polymorphisms have been associated with susceptibility to different diseases whose pathologies involve oxidative stress. In this study, we tested the hypothesis that GST polymorphisms that lead to reduced or lack of enzyme activity are associated with severe Plasmodium falciparum malarial anemia. We studied the genotypic distribution of GSTM1, GSTT1, and GSTP1 polymorphisms between mild malaria (N = 107) and severe malarial anemia (N = 50) in Tanzanian children. We did not find a significant relationship with the GSTT1 polymorphism. GSTM1-null was higher in the severe malaria anemia group but the difference was not significant (P = 0.08). However, a significant association of GSTP1 I105V genotype with severe malarial anemia was discovered (26.0% against 10.3% mild malaria, P = 0.004). We concluded that GSTP1 and possibly GSTM1 may protect against severe falciparum malaria in children.

High resistance of Plasmodium falciparum to sulphadoxine/pyrimethamine in northern Tanzania and the emergence of dhps resistance mutation at Codon 581.

BACKGROUND: Sulphadoxine-pyrimethamine (SP) a widely used treatment for uncomplicated malaria and recommended for intermittent preventive treatment of malaria in pregnancy, is being investigated for intermittent preventive treatment of malaria in infants (IPTi). High levels of drug resistance to SP have been reported from north-eastern Tanzania associated with mutations in parasite genes. This study compared the in vivo efficacy of SP in symptomatic 6-59 month children with uncomplicated malaria and in asymptomatic 2-10 month old infants. METHODOLOGY AND PRINCIPAL FINDINGS: An open label single arm (SP) standard 28 day in vivo WHO antimalarial efficacy protocol was used in 6 to 59 months old symptomatic children and a modified protocol used in 2 to 10 months old asymptomatic infants. Enrolment was stopped early (87 in the symptomatic and 25 in the asymptomatic studies) due to the high failure rate. Molecular markers were examined for recrudescence, re-infection and markers of drug resistance and a review of literature of studies looking for the 581G dhps mutation was carried out. In symptomatic children PCR-corrected early treatment failure was 38.8% (95% CI 26.8-50.8) and total failures by day 28 were 82.2% (95% CI 72.5-92.0). There was no significant difference in treatment failures between asymptomatic and symptomatic children. 96% of samples carried parasites with mutations at codons 51, 59 and 108 in the dhfr gene and 63% carried a double mutation at codons 437 and 540. 55% carried a third mutation with the addition of a mutation at codon 581 in the dhps gene. This triple: triple haplotype maybe associated with earlier treatment failure. CONCLUSION: In northern Tanzania SP is a failed drug for treatment and its utility for prophylaxis is doubtful. The study found a new combination of parasite mutations that maybe associated with increased and earlier failure. TRIAL REGISTRATION: ClinicalTrials.gov NCT00361114.

Overuse of artemisinin-combination therapy in Mto wa Mbu (river of mosquitoes), an area misinterpreted as high endemic for malaria.

BACKGROUND: Adequate malaria diagnosis and treatment remain major difficulties in rural sub-Saharan Africa. These issues deserve renewed attention in the light of first-line treatment with expensive artemisinin-combination therapy (ACT) and changing patterns of transmission intensity. This study describes diagnostic and treatment practices in Mto wa Mbu, an area that used to be hyperendemic for malaria, but where no recent assessments of transmission intensity have been conducted. METHODS: Retrospective and prospective data were collected from the two major village health clinics. The diagnosis in prospectively collected data was confirmed by microscopy. The level of transmission intensity was determined by entomological assessment and by estimating sero-conversion rates using anti-malarial antibody responses. RESULTS: Malaria transmission intensity by serological assessment was equivalent to < 1 infectious bites per person per year. Despite low transmission intensity, > 40% of outpatients attending the clinics in 2006-2007 were diagnosed with malaria. Prospective data demonstrated a very high overdiagnosis of malaria. Microscopy was unreliable with < 1% of slides regarded as malaria parasite-positive by clinic microscopists being confirmed by trained research microscopists. In addition, many 'slide negatives' received anti-malarial treatment. As a result, 99.6% (248/249) of the individuals who were treated with ACT were in fact free of malaria parasites. CONCLUSION: Transmission intensity has dropped considerably in the area of Mto wa Mbu. Despite this, most fevers are still regarded and treated as malaria, thereby ignoring true causes of febrile illness and over-prescribing ACT. The discrepancy between the perceived and actual level of transmission intensity may be present in many areas in sub-Saharan Africa and calls for greater efforts in defining levels of transmission on a local scale to help rational drug-prescribing behaviour.

An experimental hut evaluation of Olyset nets against anopheline mosquitoes after seven years use in Tanzanian villages.

BACKGROUND: Long-lasting insecticidal nets (LLINs) are advocated by WHO for protection against malaria. Of the three brands of LLINs currently approved by WHO, Olyset(R) is the only one currently granted full recommendation. With this type of LLIN, the insecticide (permethrin) is incorporated into the polyethylene fibre during manufacture and diffuses from the core to the surface, thereby maintaining surface concentrations. It has not been determined for how long Olyset nets remain protective against mosquitoes in household use. METHODS: Examples of Olyset nets, which had been in use in Tanzanian villages for seven years, were tested in experimental huts against naturally entering Anopheles gambiae and Anopheles funestus mosquitoes. Performance was compared with new Olyset nets, conventionally treated ITNs (either newly treated with alphacypermethrin or taken from local villages after 1.5 years of use) and untreated nets. All nets were artificially holed except for the seven-year Olyset nets, which had developed holes during prolonged domestic use. RESULTS: Anopheles funestus and An. gambiae in NE Tanzania are susceptible to pyrethroids. The new Olyset nets caused high mortality against An. funestus (73.9%) and An. gambiae (62.7%) in experimental huts. The seven-year Olyset nets caused 58.9% mortality against An. funestus and 40.0% mortality against An. gambiae. The freshly treated alphacypermethrin nets also caused high mortality against An. funestus (70.6%) and An. gambiae (72.0%); this decreased to 58.4% and 69.6% respectively after 1.5 years of use. The new Olyset nets inhibited blood-feeding by 40-50%. The 7 year Olyset nets showed no feeding inhibition over that shown by the untreated nets. The alphacypermethrin treated nets failed to inhibit blood-feeding after 1.5 years of use. However iHhhdn laboratory tunnel tests samples of all types of treated net including the 7 year Olyset inhibited blood-feeding by more than 95%. CONCLUSION: After seven years of use Olyset nets were still strongly insecticidal. Mosquito mortality decreased by only 20-35% over this period. However, Olyset would not provide personal protection after seven years unless it was in good condition and all holes fully repaired.

Primaquine clears submicroscopic Plasmodium falciparum gametocytes that persist after treatment with sulphadoxine-pyrimethamine and artesunate.

BACKGROUND: P. falciparum gametocytes may persist after treatment with sulphadoxine-pyrimethamine (SP) plus artesunate (AS) and contribute considerably to malaria transmission. We determined the efficacy of SP+AS plus a single dose of primaquine (PQ, 0.75 mg/kg) on clearing gametocytaemia measured by molecular methods. METHODOLOGY: The study was conducted in Mnyuzi, an area of hyperendemic malaria in north-eastern Tanzania. Children aged 3-15 years with uncomplicated P. falciparum malaria with an asexual parasite density between 500-100,000 parasites/microL were randomized to receive treatment with either SP+AS or SP+AS+PQ. P. falciparum gametocyte prevalence and density during the 42-day follow-up period were determined by real-time nucleic acid sequence-based amplification (QT-NASBA). Haemoglobin levels (Hb) were determined to address concerns about haemolysis in G6PD-deficient individuals. RESULTS: 108 individuals were randomized. Pfs25 QT-NASBA gametocyte prevalence was 88-91% at enrolment and decreased afterwards for both treatment arms. Gametocyte prevalence and density were significantly lower in children treated with SP+AS+PQ. On day 14 after treatment 3.9% (2/51) of the SP+AS+PQ treated children harboured gametocytes compared to 62.7% (32/51) of those treated with SP+AS (p<0.001). Hb levels were reduced in the week following treatment with SP+AS+PQ and this reduction was related to G6PD deficiency. The Hb levels of all patients recovered to pre-treatment levels or greater within one month after treatment. CONCLUSIONS: PQ clears submicroscopic gametocytes after treatment with SP+AS and the persisting gametocytes circulated at densities that are unlikely to contribute to malaria transmission. For individuals without severe anaemia, addition of a single dose of PQ to an efficacious antimalarial drug combination is a safe approach to reduce malaria transmission following treatment. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN61534963.

Efficacy of pyrethroid-treated nets against malaria vectors and nuisance-biting mosquitoes in Tanzania in areas with long-term insecticide-treated net use.

OBJECTIVE: To measure pyrethroid susceptibility in populations of malaria vectors and nuisance-biting mosquitoes in Tanzania and to test the biological efficacy of current insecticide formulations used for net treatment. METHODS: Anopheles gambiae Giles s.l., An. funestus Giles s.l. and Culex quinquefasciatus Say were collected during three national surveys and two insecticide-treated net (ITN) studies in Tanzania. Knockdown effect and mortality were measured in standard WHO susceptibility tests and ball-frame bio-efficacy tests. Test results from 1999 to 2004 were compared to determine trends in resistance development. RESULTS: Anopheles gambiae s.l. and An. funestus s.l. were highly susceptible to permethrin (range 87-100%) and deltamethrin (consistently 100%) in WHO tests in 1999 and 2004, while Culex quinquefasciatus susceptibility to these pyrethroids was much lower (range 7-100% and 0-84% respectively). Efficacy of pyrethroid-treated nets was similarly high against An. gambiae s.l. and An. funestus s.l. (range 82-100%) while efficacy against Cx. quinquefasciatus was considerably lower (range 2-100%). There was no indication of development of resistance in populations of An. gambiae s.l. or An. funestus s.l. where ITNs have been extensively used; however, susceptibility of nuisance-biting Cx. quinquefasciatus mosquitoes declined in some areas between 1999 and 2004. CONCLUSION: The sustained pyrethroid susceptibility of malaria vectors in Tanzania is encouraging for successful malaria control with ITNs. Continued monitoring is essential to ensure early resistance detection, particularly in areas with heavy agricultural or public health use of insecticides where resistance is likely to develop. Widespread low susceptibility of nuisance-biting Culex mosquitoes to ITNs raises concern for user acceptance of nets.

Submicroscopic Plasmodium falciparum gametocyte carriage is common in an area of low and seasonal transmission in Tanzania.

OBJECTIVE: Recently developed molecular gametocyte detection techniques have shown that submicroscopic Plasmodium falciparum gametocytes are common in symptomatic patients and can infect mosquitoes. The relevance for the infectious reservoir of malaria in the general population remains unknown. In this study, we investigated submicroscopic asexual parasitaemia and gametocytaemia in inhabitants of an area of hypoendemic and seasonal malaria in Tanzania. METHODS: Two cross-sectional malariometric surveys were conducted in the dry and wet seasons of 2005 in villages in lower Moshi, Tanzania. Finger prick blood samples were taken to determine the prevalence of P. falciparum parasites by microscopy, rapid diagnostic test and real-time nucleic acid sequence-based amplification (QT-NASBA). RESULTS: 2752 individuals participated in the surveys, of whom 1.9% (51/2721) had microscopically confirmed asexual parasites and 0.4% (10/2721) had gametocytes. In contrast, QT-NASBA revealed that 32.5% (147/453) of the individuals harboured asexual parasites and 15.0% (68/453) had gametocytes. No age dependency or seasonality was observed in submicroscopic parasite carriage. DISCUSSION: Molecular detection techniques reveal that carriage of submicroscopic asexual parasite and gametocyte densities is relatively common in this low transmission area. Submicroscopic gametocytaemia is likely to be responsible for maintaining malarial transmission in the study area.

Use of a clean delivery kit and factors associated with cord infection and puerperal sepsis in Mwanza, Tanzania.

Our objective was to determine the effectiveness of an intervention that incorporated education about the "six cleans" with the use of a clean delivery kit in preventing cord infection and puerperal sepsis. A stepped-wedge, cross-sectional study was conducted in 10 surveillance sites across two rural districts of Mwanza Region, Tanzania. A total of 3262 pregnant women between the ages of 17 and 45 years were enrolled in the study. Village health workers administered questionnaires to each mother at 5 days postpartum and inspected the infants' umbilical cord stumps for signs of infection. Newborns whose mothers used the delivery kit were 13.1 times less likely to develop cord infection than infants whose mothers did not use the kit. Furthermore, women who used the kit for delivery were 3.2 times less likely to develop puerperal sepsis than women who did not use the kit. Women who bathed before delivery were 2.6 times less likely to develop puerperal sepsis than women who did not bathe, and their infants were 3.9 times less likely to develop cord infection. Single-use delivery kits, when combined with education about clean delivery, can have a positive impact on the health of women and their newborns by significantly decreasing the likelihood of developing puerperal sepsis or cord infection.