Specific human antibody responses to Aedes aegypti and Aedes polynesiensis saliva: A new epidemiological tool to assess human exposure to disease vectors in the Pacific.

BACKGROUND: Aedes mosquitoes severely affect the health and wellbeing of human populations by transmitting infectious diseases. In French Polynesia, Aedes aegypti is the main vector of dengue, chikungunya and Zika, and Aedes polynesiensis the primary vector of Bancroftian filariasis and a secondary vector of arboviruses. Tools for assessing the risk of disease transmission or for measuring the efficacy of vector control programmes are scarce. A promising approach to quantify the human-vector contact relies on the detection and the quantification of antibodies directed against mosquito salivary proteins. METHODOLOGY/PRINCIPAL FINDINGS: An ELISA test was developed to detect and quantify the presence of immunoglobulin G (IgG) directed against proteins from salivary gland extracts (SGE) of Ae. aegypti and Ae. polynesiensis in human populations exposed to either species, through a cross-sectional study. In Tahiti and Moorea islands where Ae. aegypti and Ae. polynesiensis are present, the test revealed that 98% and 68% of individuals have developed IgG directed against Ae. aegypti and Ae. polynesiensis SGE, respectively. By comparison, ELISA tests conducted on a cohort of people from metropolitan France, not exposed to these Aedes mosquitoes, indicated that 97% of individuals had no IgG directed against SGE of either mosquito species. The analysis of additional cohorts representing different entomological Aedes contexts showed no ELISA IgG cross-reactivity between Ae. aegypti and Ae. polynesiensis SGE. CONCLUSIONS/SIGNIFICANCE: The IgG response to salivary gland extracts seems to be a valid and specific biomarker of human exposure to the bites of Ae. aegypti and Ae. polynesiensis. This new immuno-epidemiological tool will enhance our understanding of people exposure to mosquito bites, facilitate the identification of areas where disease transmission risk is high and permit to evaluate the efficacy of novel vector control strategies in Pacific islands and other tropical settings.

Evaluation of traps and lures for mosquito vectors and xenomonitoring of Wuchereria bancrofti infection in a high prevalence Samoan Village.

BACKGROUND: Elimination of lymphatic filariasis (LF) in Samoa continues to be challenging despite multiple annual mass drug campaigns aimed at stopping transmission by reducing the prevalence and density of microfilaraemia. The persistence of transmission may be partly related to the highly efficient Aedes vectors. The assessment of pathogen transmission by mosquito vectors and of vector control relies on the ability to capture mosquitoes efficiently. The aims of this study are to compare trapping methods to capture LF-infected mosquitoes and determine the role in transmission of the species of Aedes mosquitoes in the area. METHODS: Fasitoo-Tai village was the chosen site because of persistent transmission despite annual mass drug administration. Sampling methods included BioGents Sentinel (BGS) trap, human-baited collections (HBC) and the Centers for Disease Control (CDC) trap. BGS and CDC traps were baited with BG-lure, CO2, and/or octenol. Individual trap locations were geo-located and efficiency of sampling methods was evaluated using a randomized Latin-square design in two locations. Number of mosquitoes collected (male and female), as well as species for each trapping method were determined. Additionally, Ae. polynesiensis and Ae. (Finlaya) spp. females were pooled by trap method and analysed for filarial DNA. Infection prevalence was estimated using the PoolScreen software. RESULTS: The BGS trap with any type of bait collected more mosquitoes compared to both the CDC trap and the HBC. The BGS trap baited with BG-lure collected more mosquitoes than with CO2 and octenol. There were no significant differences between trapping methods in terms of proportions of infected females collected. The prevalence of filarial infection in Ae. polynesiensis and Ae. (Finlaya) spp. was estimated at 4.7% and 0.67% respectively. CONCLUSIONS: This study supports the use of the BGS trap for research on and surveillance of the mosquito vectors of LF in Samoa. The BGS trap is a suitable and safer alternative to HBC for sampling Ae. polynesiensis and Ae. (Finlaya) spp., which continue to be the predominant vectors of LF. Of concern was the high prevalence of LF in mosquitoes despite a recent mass drug administration programme. This highlights the urgency for updated policies concerning filariasis elimination in Samoa.

Monitoring and evaluation of lymphatic filariasis interventions: an improved PCR-based pool screening method for high throughput Wuchereria bancrofti detection using dried blood spots.

BACKGROUND: Effective diagnostic tools are necessary to monitor and evaluate interruption of Lymphatic Filariasis (LF) transmission. Accurate detection of Wuchereria bancrofti (Wb) microfilaria (mf) is essential to measure the impact of community treatment programmes. PCR-based assays are specific, highly sensitive tools allowing the detection of Wuchereria bancrofti DNA in human blood samples. However, current protocols describing the pool screening approach, use samples of less than 60 µl of blood, which limits the sensitivity of the pool-screen PCR assay. The purpose of this study was to improve the pool-screen PCR protocol to enhance its sensitivity and usefulness for population scale studies. FINDINGS: DNA extractions were performed with the DNeasy kit, the PCR with the Wb LDR primers and the SYBR-Green dye. Improvements of our pool-screen real-time PCR (qPCR) assay allowed the detection of as little as one Wb microfilaria diluted in a pool of at least 12 blood samples of 60 µl each. Using this assay, mf burdens can be predicted using a standard curve derived from mf spiked dried blood samples. The sensitivity achieved is equivalent to the detection of a single LF positive individual carrying a mf burden as low as 18 mf/ml, in a pool of blood samples from at least 12 individuals. CONCLUSIONS: Due to its sensitivity, rapidity and cost-effectiveness, we suggest this qPCR pool-screening assay could be used as a diagnostic tool for population- scale filariasis elimination monitoring and evaluation.

Open release of male mosquitoes infected with a wolbachia biopesticide: field performance and infection containment.

BACKGROUND: Lymphatic filariasis (LF) is a globally significant disease, with 1.3 billion persons in 83 countries at risk. A coordinated effort of administering annual macrofilaricidal prophylactics to the entire at-risk population has succeeded in impacting and eliminating LF transmission in multiple regions. However, some areas in the South Pacific are predicted to persist as transmission sites, due in part to the biology of the mosquito vector, which has led to a call for additional tools to augment drug treatments. Autocidal strategies against mosquitoes are resurging in the effort against invasive mosquitoes and vector borne disease, with examples that include field trials of genetically modified mosquitoes and Wolbachia population replacement. However, critical questions must be addressed in anticipation of full field trials, including assessments of field competitiveness of transfected males and the risk of unintended population replacement. METHODOLOGY/PRINCIPAL FINDINGS: We report the outcome of field experiments testing a strategy that employs Wolbachia as a biopesticide. The strategy is based upon Wolbachia-induced conditional sterility, known as cytoplasmic incompatibility, and the repeated release of incompatible males to suppress a population. A criticism of the Wolbachia biopesticide approach is that unintended female release or horizontal Wolbachia transmission can result in population replacement instead of suppression. We present the outcome of laboratory and field experiments assessing the competitiveness of transfected males and their ability to transmit Wolbachia via horizontal transmission. CONCLUSIONS/SIGNIFICANCE: The results demonstrate that Wolbachia-transfected Aedes polynesiensis males are competitive under field conditions during a thirty-week open release period, as indicated by mark, release, recapture and brood-hatch failure among females at the release site. Experiments demonstrate the males to be 'dead end hosts' for Wolbachia and that methods were adequate to prevent population replacement at the field site. The findings encourage the continued development and extension of a Wolbachia autocidal approach to additional medically important mosquito species.

A multicenter evaluation of diagnostic tools to define endpoints for programs to eliminate bancroftian filariasis.

Successful mass drug administration (MDA) campaigns have brought several countries near the point of Lymphatic Filariasis (LF) elimination. A diagnostic tool is needed to determine when the prevalence levels have decreased to a point that MDA campaigns can be discontinued without the threat of recrudescence. A six-country study was conducted assessing the performance of seven diagnostic tests, including tests for microfilariae (blood smear, PCR), parasite antigen (ICT, Og4C3) and antifilarial antibody (Bm14, PanLF, Urine SXP). One community survey and one school survey were performed in each country. A total of 8,513 people from the six countries participated in the study, 6,443 through community surveys and 2,070 through school surveys. Specimens from these participants were used to conduct 49,585 diagnostic tests. Each test was seen to have both positive and negative attributes, but overall, the ICT test was found to be 76% sensitive at detecting microfilaremia and 93% specific at identifying individuals negative for both microfilariae and antifilarial antibody; the Og4C3 test was 87% sensitive and 95% specific. We conclude, however, that the ICT should be the primary tool recommended for decision-making about stopping MDAs. As a point-of-care diagnostic, the ICT is relatively inexpensive, requires no laboratory equipment, has satisfactory sensitivity and specificity and can be processed in 10 minutes-qualities consistent with programmatic use. Og4C3 provides a satisfactory laboratory-based diagnostic alternative.

PCR and dissection as tools to monitor filarial infection of Aedes polynesiensis mosquitoes in French Polynesia.

BACKGROUND: Entomological methods may provide important tools for monitoring the transmission of filariasis in French Polynesia. In order to standardize our PCR method and refine our protocol to assess filarial infection levels in mosquitoes, we compared dissection of the vector, Aedes polynesiensis, with the poolscreening polymerase chain reaction (PS-PCR) assay. METHODS: (1) Mosquitoes were collected in human landing catches in five areas in Moorea island, French Polynesia. (2) A fraction of the captured mosquitoes was dissected for Wuchereria bancrofti larvae. (3) Laboratory-reared mosquitoes (uninfected as well as experimentally infected ones) were repeatedly tested to optimize a PS-PCR protocol (DNA extracts from 1-50 pooled mosquitoes were tested with an internal standardized system and primers specific for the Ssp1 repeat sequence. PCR products were analysed by gel electrophoresis). (4) Another fraction of the captured mosquitoes was assayed by PS-PCR according the optimized protocol. RESULTS: The prevalence of field-mosquito infection with W. bancrofti ranged from 1 to 8 % by dissection (L1-L3) and point estimates of infection prevalence, as assayed by PS-PCR, ranged from 0.4 to 3.7 %. There was a moderately strong correlation between larval infection rates as determined by dissection and PCR. DISCUSSION: Our results suggest that the PS-PCR assay is specific and highly sensitive for detecting parasite DNA. We obtained similar although not identical results with dissections of mosquitoes. PS-PCR appears to be adequate for testing large numbers of mosquitoes in the context of filariasis elimination programs. The role and advantages of using entomologic methods to monitor filariasis programs are discussed.

Detection and characterization of Wolbachia infections in Wuchereria bancrofti (Spirurida: Onchocercidae) var. pacifica and Aedes (Stegomyia) polynesiensis (Diptera: Culicidae).

Despite control programs based on mass drug administration (MDA) of microfilaricidal compounds, Bancroftian lymphatic filariasis remains a problem in French Polynesia. For an alternative strategy to MDA, we investigated the potential role of Wolbachia to control filarial transmission. Wolbachia are intracellular alpha-proteobacteria endosymbionts that infect a broad range of insects and nematodes. These bacteria have a suspected role in the pathogenesis of filariasis. They also may be useful in mosquito control through cytoplasmic incompatibility. To detect and characterize these bacteria in the filarial and mosquito-vectors in French Polynesia, a survey was conducted on field-collected mosquitoes and microfilariae from infected people. Samples were analyzed by a polymerase chain reaction and gene sequencing. The results indicate that these bacteria are widespread. Sequence analysis of the wsp and ftsZ genes positioned the Aedes polynesiensis Wolbachia in cluster A and Wuchereria bancrofti var. pacifica Wolbachia in cluster D. The implications for possible improved treatment and vector control are discussed.