Dataset Information

Field evaluation of DNA detection of human filarial and malaria parasites using mosquito excreta/feces.

ABSTRACT: We recently developed a superhydrophobic cone-based method for the collection of mosquito excreta/feces (E/F) for the molecular xenomonitoring of vector-borne parasites showing higher throughput compared to the traditional approach. To test its field applicability, we used this platform to detect the presence of filarial and malaria parasites in two villages of Ghana and compared results to those for detection in mosquito carcasses and human blood. We compared the molecular detection of three parasites (Wuchereria bancrofti, Plasmodium falciparum and Mansonella perstans) in mosquito E/F, mosquito carcasses and human blood collected from the same households in two villages in the Savannah Region of the country. We successfully detected the parasite DNA in mosquito E/F from indoor resting mosquitoes, including W. bancrofti which had a very low community prevalence (2.5-3.8%). Detection in the E/F samples was concordant with detection in insect whole carcasses and human blood, and a parasite not vectored by mosquitoes was detected as well.Our approach to collect and test mosquito E/F successfully detected a variety of parasites at varying prevalence in the human population under field conditions, including a pathogen (M. perstans) which is not transmitted by mosquitoes. The method shows promise for further development and applicability for the early detection and surveillance of a variety of pathogens carried in human blood.

SUBMITTER: Minetti C

PROVIDER: S-EPMC7170280 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

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Field evaluation of DNA detection of human filarial and malaria parasites using mosquito excreta/feces.

Minetti Corrado C Pilotte Nils N Zulch Michael M Canelas Tiago T Tettevi Edward J EJ Veriegh Francis B D FBD Osei-Atweneboana Mike Yaw MY Williams Steven A SA Reimer Lisa J LJ

PLoS neglected tropical diseases 20200408 4

We recently developed a superhydrophobic cone-based method for the collection of mosquito excreta/feces (E/F) for the molecular xenomonitoring of vector-borne parasites showing higher throughput compared to the traditional approach. To test its field applicability, we used this platform to detect the presence of filarial and malaria parasites in two villages of Ghana and compared results to those for detection in mosquito carcasses and human blood. We compared the molecular detection of three pa ...[more]

PMID: 32267840

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Project description:BackgroundGiven the continued successes of the world's lymphatic filariasis (LF) elimination programs and the growing successes of many malaria elimination efforts, the necessity of low cost tools and methodologies applicable to long-term disease surveillance is greater than ever before. As many countries reach the end of their LF mass drug administration programs and a growing number of countries realize unprecedented successes in their malaria intervention efforts, the need for practical molecular xenomonitoring (MX), capable of providing surveillance for disease recrudescence in settings of decreased parasite prevalence is increasingly clear. Current protocols, however, require testing of mosquitoes in pools of 25 or fewer, making high-throughput examination a challenge. The new method we present here screens the excreta/feces from hundreds of mosquitoes per pool and provides proof-of-concept for a practical alternative to traditional methodologies resulting in significant cost and labor savings.Methodology/principal findingsExcreta/feces of laboratory reared Aedes aegypti or Anopheles stephensi mosquitoes provided with a Brugia malayi microfilaria-positive or Plasmodium vivax-positive blood meal respectively were tested for the presence of parasite DNA using real-time PCR. A titration of samples containing various volumes of B. malayi-negative mosquito feces mixed with positive excreta/feces was also tested to determine sensitivity of detection. Real-time PCR amplification of B. malayi and P. vivax DNA from the excreta/feces of infected mosquitoes was demonstrated, and B. malayi DNA in excreta/feces from one to two mf-positive blood meal-receiving mosquitoes was detected when pooled with volumes of feces from as many as 500 uninfected mosquitoes.Conclusions/significanceWhile the operationalizing of excreta/feces testing may require the development of new strategies for sample collection, the high-throughput nature of this new methodology has the potential to greatly reduce MX costs. This will prove particularly useful in post-transmission-interruption settings, where this inexpensive approach to long-term surveillance will help to stretch the budgets of LF and malaria elimination programs. Furthermore, as this methodology is adaptable to the detection of both single celled (P. vivax) and multicellular eukaryotic pathogens (B. malayi), exploration of its use for the detection of various other mosquito-borne diseases including viruses should be considered. Additionally, integration strategies utilizing excreta/feces testing for the simultaneous surveillance of multiple diseases should be explored.

Project description:Background: Results from an increasing number of studies suggest that mosquito excreta/feces (E/F) testing has considerable potential to serve as a supplement for traditional molecular xenomonitoring techniques. However, as the catalogue of possible use-cases for this methodology expands, and the list of amenable pathogens grows, a number of fundamental methods-based questions remain. Answering these questions is critical to maximizing the utility of this approach and to facilitating its successful implementation as an effective tool for molecular xenomonitoring. Methods: Utilizing E/F produced by mosquitoes or tsetse flies experimentally exposed to Brugia malayi, Plasmodium falciparum, or Trypanosoma brucei brucei, factors such as limits of detection, throughput of testing, adaptability to use with competent and incompetent vector species, and effects of additional blood feedings post parasite-exposure were evaluated. Two platforms for the detection of pathogen signal (quantitative real-time PCR and digital PCR (dPCR)) were also compared, with strengths and weaknesses examined for each. Results: Experimental results indicated that high throughput testing is possible when evaluating mosquito E/F for the presence of either B. malayi or P. falciparum from both competent and incompetent vector mosquito species. Furthermore, following exposure to pathogen, providing mosquitoes with a second, uninfected bloodmeal did not expand the temporal window for E/F collection during which pathogen detection was possible. However, this collection window did appear longer in E/F collected from tsetse flies following exposure to T. b. brucei. Testing also suggested that dPCR may facilitate detection through its increased sensitivity. Unfortunately, logistical obstacles will likely make the large-scale use of dPCR impractical for this purpose. Conclusions: By examining many E/F testing variables, expansion of this technology to a field-ready platform has become increasingly feasible. However, translation of this methodology from the lab to the field will first require field-based pilot studies aimed at assessing the efficacy of E/F screening.

Project description:Traditional screening for arboviruses in mosquitoes requires a priori knowledge and the utilization of appropriate assays for their detection. Mosquitoes can also provide other valuable information, including unexpected or novel arboviruses, nonarboviral pathogens ingested from hosts they feed on, and their own genetic material. Metagenomic analysis using next-generation sequencing (NGS) is a rapidly advancing technology that allows us to potentially obtain all this information from a mosquito sample without any prior knowledge of virus, host, or vector. Moreover, it has been recently demonstrated that pathogens, including arboviruses and parasites, can be detected in mosquito excreta by molecular methods. In this study, we investigated whether RNA viruses could be detected in mosquito excreta by NGS. Excreta samples were collected from Aedes vigilax and Culex annulirostris experimentally exposed to either Ross River or West Nile viruses and from field mosquitoes collected across Queensland, Australia. Total RNA was extracted from the excreta samples, reverse transcribed to cDNA, and sequenced using the Illumina NextSeq 500 platform. Bioinformatic analyses from the generated reads demonstrate that mosquito excreta provide sufficient RNA for NGS, allowing the assembly of near-full-length viral genomes. We detected Australian Anopheles totivirus, Wuhan insect virus 33, and Hubei odonate virus 5 and identified seven potentially novel viruses closely related to members of the order Picornavirales (2/7) and to previously described, but unclassified, RNA viruses (5/7). Our results suggest that metagenomic analysis of mosquito excreta has great potential for virus discovery and for unbiased arbovirus surveillance in the near future.IMPORTANCE When a mosquito feeds on a host, it ingests not only its blood meal but also an assortment of microorganisms that are present in the blood, thus acting as an environmental sampler. By using specific tests, it is possible to detect arthropod-borne viruses (arboviruses) like dengue and West Nile viruses in mosquito excreta. Here, we explored the use of next-generation sequencing (NGS) for unbiased detection of RNA viruses present in excreta from experimentally infected and field-collected mosquitoes. We have demonstrated that mosquito excreta provide a suitable template for NGS and that it is possible to recover and assemble near-full-length genomes of both arboviruses and insect-borne viruses, including potentially novel ones. These results importantly show the direct practicality of the use of mosquito excreta for NGS, which in the future could be used for virus discovery, environmental virome sampling, and arbovirus surveillance.

Project description:BackgroundThere is growing interest in the potential to modify houses to target mosquitoes with insecticides or repellents as they search for human hosts. One version of this 'Lethal House Lure' approach is the In2Care® EaveTube, which consists of a section of polyvinyl chloride (PVC) pipe fitted into a closed eave, with an insert comprising electrostatic netting treated with insecticide powder placed inside the tube. Preliminary evidence suggests that when combined with screening of doors and windows, there is a reduction in entry of mosquitoes and an increase in mortality. However, the rate of overnight mortality remains unclear. The current study used a field enclosure built around experimental huts to investigate the mortality of cohorts of mosquitoes over multiple nights.MethodsAnopheles gambiae sensu lato mosquitoes were collected from the field as larvae and reared through to adult. Three-to-five days old adult females were released inside an enclosure housing two modified West African style experimental huts at a field site in M'be, Côte d'Ivoire. Huts were either equipped with insecticide-treated tubes at eave height and had closed windows (treatment) or had open windows and open tubes (controls). The number of host-seeking mosquitoes entering the huts and cumulative mortality were monitored over 2 or 4 days.ResultsVery few (0-0.4%) mosquitoes were able to enter huts fitted with insecticide-treated tubes and closed windows. In contrast, mosquitoes continually entered the control huts, with a cumulative mean of 50-80% over 2 to 4 days. Baseline mortality with control huts was approximately 2-4% per day, but the addition of insecticide-treated tubes increased mortality to around 25% per day. Overall cumulative mortality was estimated to be up to 87% over 4 days when huts were fitted with tubes.ConclusionOnly 20-25% of mosquitoes contacted insecticide-treated tubes or entered control huts in a given night. However, mosquitoes continue to host search over sequential nights, and this can lead to high cumulative mortality over 2 to 4 days. This mortality should contribute to community-level reduction in transmission assuming sufficient coverage of the intervention.

Dataset Information

Field evaluation of DNA detection of human filarial and malaria parasites using mosquito excreta/feces.

Publications

Field evaluation of DNA detection of human filarial and malaria parasites using mosquito excreta/feces.

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