Edward K. Thomsen

Insecticidal bed nets and filariasis transmission in Papua New Guinea.

BACKGROUND Global efforts to eliminate lymphatic filariasis are based on the annual mass administration of antifilarial drugs to reduce the microfilaria reservoir available to the mosquito vector. Insecticide-treated bed nets are being widely used in areas in which filariasis and malaria are coendemic. METHODS We studied five villages in which five annual mass administrations of antifilarial drugs, which were completed in 1998, reduced the transmission of Wuchereria bancrofti, one of the nematodes that cause lymphatic filariasis. A total of 21,899 anopheles mosquitoes were collected for 26 months before and 11 to 36 months after bed nets treated with long-lasting insecticide were distributed in 2009. We evaluated the status of filarial infection and the presence of W. bancrofti DNA in anopheline mosquitoes before and after the introduction of insecticide-treated bed nets. We then used a model of population dynamics to estimate the probabilities of transmission cessation. RESULTS Village-specific rates of bites from anopheline mosquitoes ranged from 6.4 to 61.3 bites per person per day before the bed-net distribution and from 1.1 to 9.4 bites for 11 months after distribution (P<0.001). During the same period, the rate of detection of W. bancrofti in anopheline mosquitoes decreased from 1.8% to 0.4% (P=0.005), and the rate of detection of filarial DNA decreased from 19.4% to 14.9% (P=0.13). The annual transmission potential was 5 to 325 infective larvae inoculated per person per year before the bed-net distribution and 0 after the distribution. Among all five villages with a prevalence of microfilariae of 2 to 38%, the probability of transmission cessation increased from less than 1.0% before the bed-net distribution to a range of 4.9 to 95% in the 11 months after distribution. CONCLUSIONS Vector control with insecticide-treated bed nets is a valuable tool for W. bancrofti elimination in areas in which anopheline mosquitoes transmit the parasite. (Funded by the U.S. Public Health Service and the National Institutes of Health.).

Efficacy, Safety, and Pharmacokinetics of Coadministered Diethylcarbamazine, Albendazole, and Ivermectin for Treatment of Bancroftian Filariasis.

BACKGROUND Available treatments for lymphatic filariasis (LF) are limited in their longterm clearance of microfilaria from the blood. The safety and efficacy of a single-dose triple-drug therapy of the antifilarial drugs diethylcarbamazine (DEC), ivermectin (IVM), and albendazole (ALB) for LF are unknown. METHODS We performed a pilot study to test the efficacy, safety, and pharmacokinetics of single-dose DEC, IVM, and ALB in Wuchereria bancrofti-infected Papua New Guineans. Adults were randomized into 2 treatment arms, DEC 6 mg/kg + ALB 400 mg (N = 12) or DEC 6 mg/kg + ALB 400 mg + IVM 200 μg/kg (N = 12), and monitored for microfilaria, parasite antigenemia, adverse events (AEs), and serum drug levels. RESULTS Triple-drug therapy induced >2-log reductions in microfilaria levels at 36 and 168 hours after treatment compared with approximately 1-log reduction with 2 drugs. All 12 individuals who received 3 drugs were microfilaria negative 1 year after treatment, whereas 11 of 12 individuals in the 2-drug regimen were microfilaria positive. In 6 participants followed 2 years after treatment, those who received 3 drugs remained microfilaria negative. AEs, particularly fever, myalgias, pruritus, and proteinuria/hematuria, occurred in 83% vs 50% of those receiving triple-drug compared to 2-drug treatment respectively (P = .021); all resolved within 7 days after treatment. No serious AEs were observed in either group. There was no significant effect of IVM on DEC or ALB drug levels. CONCLUSIONS Triple-drug therapy is safe and more effective than DEC + ALB for Bancroftian filariasis and has the potential to accelerate elimination of lymphatic filariasis. CLINICAL TRIALS REGISTRATION NCT01975441.

Underpinning sustainable vector control through informed insecticide resistance management.

Background There has been rapid scale-up of malaria vector control in the last ten years. Both of the primary control strategies, long-lasting pyrethroid treated nets and indoor residual spraying, rely on the use of a limited number of insecticides. Insecticide resistance, as measured by bioassay, has rapidly increased in prevalence and has come to the forefront as an issue that needs to be addressed to maintain the sustainability of malaria control and the drive to elimination. Zambias programme reported high levels of resistance to the insecticides it used in 2010, and, as a result, increased its investment in resistance monitoring to support informed resistance management decisions. Methodology/Principal Findings A country-wide survey on insecticide resistance in Zambian malaria vectors was performed using WHO bioassays to detect resistant phenotypes. Molecular techniques were used to detect target-site mutations and microarray to detect metabolic resistance mechanisms. Anopheles gambiae s.s. was resistant to pyrethroids, DDT and carbamates, with potential organophosphate resistance in one population. The resistant phenotypes were conferred by both target-site and metabolic mechanisms. Anopheles funestus s.s. was largely resistant to pyrethroids and carbamates, with potential resistance to DDT in two locations. The resistant phenotypes were conferred by elevated levels of cytochrome p450s. Conclusions/Significance Currently, the Zambia National Malaria Control Centre is using these results to inform their vector control strategy. The methods employed here can serve as a template to all malaria-endemic countries striving to create a sustainable insecticide resistance management plan.

Molecular-Based Assay for Simultaneous Detection of Four Plasmodium spp. and Wuchereria bancrofti Infections

Four major malaria-causing Plasmodium spp. and lymphatic filariasis-causing Wuchereria bancrofti are co-endemic in many tropical and sub-tropical regions. Among molecular diagnostic assays, multiplex polymerase chain reaction (PCR)-based assays for the simultaneous detection of DNAs from these parasite species are currently available only for P. falciparum and W. bancrofti or P. vivax and W. bancrofti. Using a post-PCR oligonucleotide ligation detection reaction-fluorescent microsphere assay (LDR-FMA), we developed a multiplex assay that has the capability to simultaneously detect all four Plasmodium spp. and W. bancrofti infections in blood samples. Compared with microfilarial positivity in the blood, the LDR-FMA assay is highly concordant (91%), sensitive (86%), and specific (94%), and has high reproducibility for Plasmodium spp. (85-93%) and W. bancrofti (90%) diagnoses. The development of this assay for the simultaneous diagnosis of multiple parasitic infections enables efficient screening of large numbers of human blood and mosquito samples from co-endemic areas.

Mosquito-Parasite Interactions Can Shape Filariasis Transmission Dynamics and Impact Elimination Programs

The relationship between mosquito vectors and lymphatic filariasis (LF) parasites can result in a range of transmission outcomes. Anophelines are generally characterized as poor vectors due to an inability to support development at low densities. However, it is important to understand the potential for transmission in natural vectors to maximize the success of elimination efforts. Primary vectors in Papua New Guinea (n = 1209) were dissected following exposure to microfilaremic blood (range 8–233 mf/20 µl). We examined density dependent and species-specific parasite prevalence, intensity and yield, barriers to parasite development as well as impacts on mosquito survival. We observed strikingly different parasite prevalence and yield among closely related species. Prevalence of infective stage larvae (L3s) ranged from 4.2% to 23.7% in An. punctulatus, 24.5% to 68.6% in An. farauti s.s. and 61.9% to 100% in An. hinesorum at low and high density exposures, respectively. Injection experiments revealed the greatest barrier to parasite development involved passage from the midgut into the hemocoel. The ratio of L3 to ingested mf at low densities was higher in An. hinesorum (yield = 1.0) and An. farauti s.s. (yield = 0.5) than has been reported in other anopheline vectors. There was a negative relationship between mosquito survival and bloodmeal mf density. In An. farauti s.s., increased parasite yield and survival at low densities suggest greater competence at low microfilaremias. In Papua New Guinea the likelihood of transmission will be strongly influenced by vector composition and changes in the mf reservoir as a result of elimination efforts. Global elimination efforts will be strengthened by the knowledge of transmission potential in the context of current control measures.

High Throughput Multiplex Assay for Species Identification of Papua New Guinea Malaria Vectors: Members of the Anopheles punctulatus (Diptera: Culicidae) Species Group

Malaria and filariasis are transmitted in the Southwest Pacific region by Anopheles punctulatus sibling species including An. punctulatus, An. koliensis, the An. farauti complex 1-8 (includes An. hinesorum [An. farauti 2], An. torresiensis [An. farauti 3]). Distinguishing these species from each other requires molecular diagnostic methods. We developed a multiplex polymerase chain reaction (PCR)-based assay specific for known species-specific nucleotide differences in the internal transcribed spacer 2 region and identified the five species most frequently implicated in transmitting disease (An. punctulatus, An. koliensis, An. farauti 1, An. hinesorum, and An. farauti 4). A set of 340 individual mosquitoes obtained from seven Papua New Guinea provinces representing a variety of habitats were analyzed by using this multiplex assay. Concordance between molecular and morphological diagnosis was 56.4% for An. punctulatus, 85.3% for An. koliensis, and 88.9% for An. farauti. Among 158 mosquitoes morphologically designated as An. farauti, 33 were re-classified by PCR as An. punctulatus, 4 as An. koliensis, 26 as An. farauti 1, 49 as An. hinesorum, and 46 as An. farauti 4. Misclassification results from variable coloration of the proboscis and overlap of An. punctulatus, An. koliensis, the An. farauti 4. This multiplex technology enables further mosquito strain identification and simultaneous detection of microbial pathogens.

Pyrethroid susceptibility in natural populations of the Anopheles punctulatus group (Diptera: Culicidae) in Papua New Guinea.

The development of insecticide resistance has compromised mosquito control efforts in many parts of the world. Papua New Guinea (PNG) has a long history of dichlorodiphenyltrichloroethane (DDT) use and currently distributes pyrethroid-treated nets for malaria control. This study is the first to investigate the status of pyrethroid resistance in the Anopheles punctulatus group, the major malaria and filariasis vectors of PNG. The study used World Health Organization standard susceptibility bioassays to detect knockdown phenotypes and a novel nested polymerase chain reaction to detect the knockdown resistant (kdr) allele in these vectors. Our results show 100% susceptibility to pyrethroids in all populations surveyed and an absence of the kdr allele.

An Operational Framework for Insecticide Resistance Management Planning

Country-wide planning and coordination can improve sustainability of vectorborne disease control.

Multiplex assay for species identification and monitoring of insecticide resistance in Anopheles punctulatus group populations of Papua New Guinea.

Anopheles punctulatus sibling species (An. punctulatus s.s., Anopheles koliensis, and Anopheles farauti species complex [eight cryptic species]) are principal vectors of malaria and filariasis in the Southwest Pacific. Given significant effort to reduce malaria and filariasis transmission through insecticide-treated net distribution in the region, effective strategies to monitor evolution of insecticide resistance among An. punctulatus sibling species is essential. Mutations in the voltage-gated sodium channel (VGSC) gene have been associated with knock-down resistance (kdr) to pyrethroids and DDT in malarious regions. By examining VGSC sequence polymorphism we developed a multiplex assay to differentiate wild-type versus kdr alleles and query intron-based polymorphisms that enable simultaneous species identification. A survey including mosquitoes from seven Papua New Guinea Provinces detected no kdr alleles in any An. punctulatus species. Absence of VGSC sequence introgression between species and evidence of geographic separation within species suggests that kdr must be monitored in each An. punctulatus species independently.

Mosquito behaviour change after distribution of bednets results in decreased protection against malaria exposure

Abstract Background. Behavioral resilience in mosquitoes poses a significant challenge to mosquito control. Although behavior changes in anopheline vectors have been reported over the last decade, there are no empirical data to suggest they compromise the efficacy of vector control in reducing malaria transmission. Methods. In this study, we quantified human exposure to both bites and infective bites of a major malaria vector in Papua New Guinea over the course of 4 years surrounding nationwide bednet distribution. We also quantified malaria infection prevalence in the human population during the same time period. Results. We observed a shift in mosquito biting to earlier hours of the evening, before individuals are indoors and protected by bednets, followed by a return to preintervention biting rates. As a result, net users and non–net users experienced higher levels of transmission than before the intervention. The personal protection provided by a bednet decreased over the study period and was lowest in the adult population, who may be an important reservoir for transmission. Malaria prevalence decreased in only 1 of 3 study villages after the distribution. Discussion. This study highlights the necessity of validating and deploying vector control measures targeting outdoor exposure to control and eliminate malaria.