Nationwide Inventory of Mosquitoes and the Distribution of Invasive Aedes (Stegomyia) albopictus (Skuse, 1894) on the Islands of Sao Tome and Principe in Central Africa.

Aedes albopictus (Skuse, 1894), a mosquito originating in Asia, has been introduced to Africa since the 2000s. The mosquito is not only a nuisance but is capable of transmitting various arboviruses. The current study summarized our entomological surveys in the Democratic Republic of Sao Tome and Principe during 2000 to 2016. Adult mosquitoes were collected by sweep nets, human landing catches, and Centers for Disease Control (CDC) light traps, and the immatures were collected from water-filled habitats at 15 sentinel sites and reared to adulthood. Species identification was performed based on morphologic characteristics. Fragments of the cytochrome C oxidase subunit I (COI) and the Wolbachia surface protein (wsp) genes were amplified for mosquitoes collected in Principe. New records of four mosquito species were reported. Aedes albopictus was identified in 2015. The larvae were found distributed over the nation and were predominately in artificial water-holding containers (488/2698, 18.1%). The highest positive rate was observed in used tires in Príncipe (114/250, 45.6%). Mitochondrial DNA analysis revealed low genetic diversity among the invasive populations, but all tested specimens were superinfected by Wolbachia. The ability of Ae. albopictus to adapt to new environments and its involvement in disease transmission make the surveillance and control of this species particularly important.

Genetic surveillance reveals low, sustained malaria transmission with clonal replacement in Sao Tome and Principe.

Despite efforts to eliminate malaria in Sao Tome and Principe (STP), cases have recently increased. Understanding residual transmission structure is crucial for developing effective elimination strategies. This study collected surveillance data and generated amplicon sequencing data from 980 samples between 2010 and 2016 to examine the genetic structure of the parasite population. The mean multiplicity of infection (MOI) was 1.3, with 11% polyclonal infections, indicating low transmission intensity. Temporal trends of these genetic metrics did not align with incidence rates, suggesting that changes in genetic metrics may not straightforwardly reflect changes in transmission intensity, particularly in low transmission settings where genetic drift and importation have a substantial impact. While 88% of samples were genetically linked, continuous turnover in genetic clusters and changes in drug-resistance haplotypes were observed. Principal component analysis revealed some STP samples were genetically similar to those from Central and West Africa, indicating possible importation. These findings highlight the need to prioritize several interventions such as targeted interventions against transmission hotspots, reactive case detection, and strategies to reduce the introduction of new parasites into this island nation as it approaches elimination. This study also serves as a case study for implementing genetic surveillance in a low transmission setting.

Dynamic changes in genetic diversity, drug resistance mutations, and treatment outcomes of falciparum malaria from the low-transmission to the pre-elimination phase on the islands of São Tomé and Príncipe.

BACKGROUND: With effective vector control and case management, substantial progress has been made towards eliminating malaria on the islands of São Tomé and Príncipe (STP). This study assessed the dynamic changes in the genetic diversity of Plasmodium falciparum, the anti-malarial drug resistance mutations, and malaria treatment outcomes between 2010 and 2016 to provide insights for the prevention of malaria rebounding. METHODS: Polymorphic regions of merozoite surface proteins 1 and 2 (msp1 and msp2) were sequenced in 118 dried blood spots (DBSs) collected from malaria patients who had visited the Central Hospital in 2010-2016. Mutations in the multi-drug resistance I (pfmdr1), chloroquine resistance transporter (pfcrt), and kelch 13 (pfk13) genes were analysed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequencing in 111 DBSs. A total of 7482 cases that completed a 28-day follow-up were evaluated for treatment outcomes based on the microscopic results. Regression models were used to characterize factors associated with levels of parasite density and treatment failures. RESULTS: Parasite strains in STP showed significant changes during and after the peak incidence in 2012. The prevalent allelic type in msp1 changed from K1 to MAD20, and that in msp2 changed from 3D7/IC to FC27. The dominant alleles of drug-resistance markers were pfmdr1 86Y, 184F, D1246, and pfcrt 76 T (Y-F-D-T, 51.4%). The average parasite density in malaria cases declined threefold from low-transmission (2010-2013) to pre-elimination period (2014-2016). Logistic regression models showed that patients with younger age (OR for age = 0.97-0.98, p < 0.001), higher initial parasite density (log10-transformed, OR = 1.44, p < 0.001), and receiving quinine treatment (compared to artemisinin-based combination therapy, OR = 1.91-1.96, p < 0.001) were more likely to experience treatment failures during follow-up. CONCLUSIONS: Plasmodium falciparum in STP had experienced changes in prevalent strains, and increased mutation frequencies in drug-resistance genes from the low-transmission to the pre-elimination settings. Notably, patients with younger age and receiving quinine treatment were more likely to show parasitological treatment failure during follow-up. Therapeutic efficacy should be carefully monitored to inform future treatment policy in STP.

Effects of indoor residual spraying and outdoor larval control on Anopheles coluzzii from São Tomé and Príncipe, two islands with pre-eliminated malaria.

BACKGROUND: Vector control is a key component of malaria prevention. Two major vector control strategies have been implemented in São Tomé and Príncipe (STP), indoor residual spraying (IRS) and outdoor larval control using Bacillus thuringiensis israelensis (Bti). This study evaluated post-intervention effects of control strategies on vector population density, composition, and knockdown resistance mutation, and their implications for malaria epidemiology in STP. METHODS: Mosquitoes were collected by indoor and outdoor human landing catches and mosquito light traps in seven districts. Mosquito density was calculated by numbers of captured adult mosquitoes/house/working hour. Mitochondrial cytochrome c oxidase subunit I (COI) was PCR amplified and sequenced to understand the spatial-temporal population composition of malaria vector in STP. Knockdown resistance L1014F mutation was detected using allele-specific PCR. To estimate the malaria transmission risks in STP, a negative binomial regression model was constructed. The response variable was monthly incidence, and the explanatory variables were area, rainfall, entomological inoculation rate (EIR), and kdr mutation frequency. RESULTS: Malaria vector in STP is exophilic Anopheles coluzzii with significant population differentiation between Príncipe and São Tomé (mean FST = 0.16, p < 0.001). Both vector genetic diversity and knockdown resistance mutation were relatively low in Príncipe (mean of kdr frequency = 15.82%) compared to São Tomé (mean of kdr frequency = 44.77%). Annual malaria incidence rate in STP had been rapidly controlled from 37 to 2.1% by three rounds of country-wide IRS from 2004 to 2007. Long-term application of Bti since 2007 kept the mosquito density under 10 mosquitoes/house/hr/month, and malaria incidence rate under 5% after 2008, except for a rising that occurred in 2012 (incidence rate = 6.9%). Risk factors of area (São Tomé compared to Príncipe), rainfall, outdoor EIR, and kdr mutation frequency could significantly increase malaria incidence by 9.33-11.50, 1.25, 1.07, and 1.06 fold, respectively. CONCLUSIONS: Indoor residual spraying could rapidly decrease Anopheles density and malaria incidence in STP. Outdoor larval control using Bti is a sustainable approach for controlling local vector with exophilic feature and insecticide resistance problem. Vector control interventions should be intensified especially at the north-eastern part of São Tomé to minimize impacts of outbreaks.

Rapid control of malaria by means of indoor residual spraying of alphacypermethrin in the Democratic Republic of São Tomé and Príncipe.

A nationwide yearly cycle of indoor residual spraying (IRS) with a pyrethroid, alphacypermethrin, at a dosage of 50 mg/m(2) was instituted in 2004 in the Democratic Republic of São Tomé and Príncipe. Rates of IRS acceptance were high, varying from 82% to 95% for dwellings and outhouses. Epidemiologic surveys of the children < 9 years of age before and after the first IRS cycle revealed a rapid reduction in malaria. Overall prevalence of malaria parasitemia for all districts was lowered from 20.1% to 2.8% at 12 months after the first IRS and reached 0.7% at 8 months after the second IRS. Longer insecticidal persistence was found on wood than on cement with alphacypermethrin.