Windborne long-distance migration of malaria mosquitoes in the Sahel.

Over the past two decades efforts to control malaria have halved the number of cases globally, yet burdens remain high in much of Africa and the elimination of malaria has not been achieved even in areas where extreme reductions have been sustained, such as South Africa1,2. Studies seeking to understand the paradoxical persistence of malaria in areas in which surface water is absent for 3-8 months of the year have suggested that some species of Anopheles mosquito use long-distance migration3. Here we confirm this hypothesis through aerial sampling of mosquitoes at 40-290 m above ground level and provide-to our knowledge-the first evidence of windborne migration of African malaria vectors, and consequently of the pathogens that they transmit. Ten species, including the primary malaria vector Anopheles coluzzii, were identified among 235 anopheline mosquitoes that were captured during 617 nocturnal aerial collections in the Sahel of Mali. Notably, females accounted for more than 80% of all of the mosquitoes that we collected. Of these, 90% had taken a blood meal before their migration, which implies that pathogens are probably transported over long distances by migrating females. The likelihood of capturing Anopheles species increased with altitude (the height of the sampling panel above ground level) and during the wet seasons, but variation between years and localities was minimal. Simulated trajectories of mosquito flights indicated that there would be mean nightly displacements of up to 300 km for 9-h flight durations. Annually, the estimated numbers of mosquitoes at altitude that cross a 100-km line perpendicular to the prevailing wind direction included 81,000 Anopheles gambiae sensu stricto, 6 million A. coluzzii and 44 million Anopheles squamosus. These results provide compelling evidence that millions of malaria vectors that have previously fed on blood frequently migrate over hundreds of kilometres, and thus almost certainly spread malaria over these distances. The successful elimination of malaria may therefore depend on whether the sources of migrant vectors can be identified and controlled.

Tracking SARS-CoV-2 seropositivity in rural communities using blood-fed mosquitoes.

The spread of SARS-CoV-2 cannot be well monitored and understood in areas without capacity for effective disease surveillance. Countries with a young population will have disproportionately large numbers of asymptomatic or pauci-symptomatic infections, further hindering detection of infection in the population. Sero-surveillance on a country-wide scale by trained medical professionals may be limited in scope in resource limited setting such as Mali. Novel ways of broadly sampling the human population in a non-invasive method would allow for large-scale surveillance at a reduced cost. Here we evaluate the collection of naturally bloodfed mosquitoes to test for human anti-SARS-CoV-2 antibodies in the laboratory and at five field locations in Mali. Immunoglobulin-G antibodies were found to be readily detectable within the mosquito bloodmeals by a bead-based immunoassay at least through 10 hours post-feeding with high sensitivity (0.900 ± 0.059) and specificity (0.924 ± 0.080), respectively, indicating that most blood-fed mosquitoes collected indoors during early morning hours (and thus, have likely fed the previous night) are viable samples for analysis. We find that reactivity to four SARS-CoV-2 antigens rose during the pandemic from pre-pandemic levels. Consistent with other sero-surveillance studies in Mali, crude seropositivity of blood sampled via mosquitoes was 6.3% in October/November 2020 over all sites, and increased to 25.1% overall, with the town closest to Bamako reaching 46.7% in February of 2021. Mosquito bloodmeals a viable target for conventional immunoassays, and therefore country-wide sero-surveillance of human diseases (both vector-borne and non-vector-borne) is attainable in areas where human-biting mosquitoes are common, and is an informative, cost-effective, non-invasive sampling option.

Tracking SARS-CoV-2 seropositivity in rural communities using blood-fed mosquitoes: a proof-of-concept study.

Background: The spread of SARS-CoV-2 cannot be well monitored and understood in areas without capacity for effective disease surveillance. Countries with a young population will have disproportionately large numbers of asymptomatic or pauci-symptomatic infections, further hindering detection of infection. Sero-surveillance on a country-wide scale by trained medical professionals may be limited in a resource-limited setting such as Mali. Novel ways of broadly sampling the human population in a non-invasive method would allow for large-scale surveillance at a reduced cost. Approach: Here we evaluate the collection of naturally blood-fed mosquitoes to test for human anti-SARS-CoV-2 antibodies in the laboratory and at five field locations in Mali. Results: Immunoglobulin-G antibodies to multiple SARS-CoV-2 antigens were readily detected in mosquito bloodmeals by bead-based immunoassay through at least 10 h after feeding [mean sensitivity of 0.92 (95% CI 0.78-1) and mean specificity of 0.98 (95% CI 0.88-1)], indicating that most blood-fed mosquitoes collected indoors during early morning hours (and likely to have fed the previous night) are viable samples for analysis. We found that reactivity to four SARS-CoV-2 antigens rose during the pandemic from pre-pandemic levels. The crude seropositivity of blood sampled via mosquitoes was 6.3% in October and November 2020 across all sites, and increased to 25.1% overall by February 2021, with the most urban site reaching 46.7%, consistent with independent venous blood-based sero-surveillance estimates. Conclusions: We have demonstrated that using mosquito bloodmeals, country-wide sero-surveillance of human diseases (both vector-borne and non-vector-borne) is possible in areas where human-biting mosquitoes are common, offering an informative, cost-effective, and non-invasive sampling option.

Isotopic evidence that aestivation allows malaria mosquitoes to persist through the dry season in the Sahel.

Data suggest that the malaria vector mosquito Anopheles coluzzii persists during the dry season in the Sahel through a dormancy mechanism known as aestivation; however, the contribution of aestivation compared with alternative strategies such as migration is unknown. Here we marked larval Anopheles mosquitoes in two Sahelian villages in Mali using deuterium (2H) to assess the contribution of aestivation to persistence of mosquitoes through the seven-month dry season. After an initial enrichment period, 33% of An. coluzzii mosquitoes were strongly marked. Seven months following enrichment, multiple analysis methods supported the ongoing presence of marked mosquitoes, compatible with the prediction that the fraction of marked mosquitoes should remain stable throughout the dry season if local aestivation is occurring. The results suggest that aestivation is a major persistence mechanism of An. coluzzii in the Sahel, contributing at least 20% of the adults at the onset of rains. This persistence strategy could influence mosquito control and malaria elimination campaigns.

Diversity, composition, altitude, and seasonality of high-altitude windborne migrating mosquitoes in the Sahel: Implications for disease transmission.

Recent studies have reported Anopheles mosquitoes captured at high-altitude (40-290 m above ground) in the Sahel. Here, we describe this migration modality across genera and species of African Culicidae and examine its implications for disease transmission and control. As well as Anopheles, six other genera-Culex, Aedes, Mansonia, Mimomyia, Lutzia, and Eretmapodites comprised 90% of the 2,340 mosquitoes captured at altitude. Of the 50 molecularly confirmed species (N = 2,107), 33 species represented by multiple specimens were conservatively considered high-altitude windborne migrants, suggesting it is a common migration modality in mosquitoes (31-47% of the known species in Mali), and especially in Culex (45-59%). Overall species abundance varied between 2 and 710 specimens/species (in Ae. vittatus and Cx. perexiguus, respectively). At altitude, females outnumbered males 6:1, and 93% of the females have taken at least one blood meal on a vertebrate host prior to their departure. Most taxa were more common at higher sampling altitudes, indicating that total abundance and diversity are underestimated. High-altitude flight activity was concentrated between June and November coinciding with availability of surface waters and peak disease transmission by mosquitoes. These hallmarks of windborne mosquito migration bolster their role as carriers of mosquito-borne pathogens (MBPs). Screening 921 mosquitoes using pan-Plasmodium assays revealed that thoracic infection rate in these high-altitude migrants was 2.4%, providing a proof of concept that vertebrate pathogens are transported by windborne mosquitoes at altitude. Fourteen of the 33 windborne mosquito species had been reported as vectors to 25 MBPs in West Africa, which represent 32% of the MBPs known in that region and include those that inflict the heaviest burden on human and animal health, such as malaria, yellow fever, dengue, and Rift Valley fever. We highlight five arboviruses that are most likely affected by windborne mosquitoes in West Africa: Rift Valley fever, O'nyong'nyong, Ngari, Pangola, and Ndumu. We conclude that the study of windborne spread of diseases by migrating insects and the development of surveillance to map the sources, routes, and destinations of vectors and pathogens is key to understand, predict, and mitigate existing and new threats of public health.

The Effects of High-Altitude Windborne Migration on Survival, Oviposition, and Blood-Feeding of the African Malaria Mosquito, Anopheles gambiae s.l. (Diptera: Culicidae).

Recent results of high-altitude windborne mosquito migration raised questions about the viability of these mosquitoes despite ample evidence that many insect species, including other dipterans, have been known to migrate regularly over tens or hundreds of kilometers on high-altitude winds and retain their viability. To address these concerns, we subjected wild Anopheles gambiae s.l. Giles mosquitoes to a high-altitude survival assay, followed by oviposition (egg laying) and blood feeding assays. Despite carrying out the survival assay under exceptionally harsh conditions that probably provide the lowest survival potential following high altitude flight, a high proportion of the mosquitoes survived for 6- and even 11-h assay durations at 120- to 250-m altitudes. Minimal differences in egg laying success were noted between mosquitoes exposed to high altitude survival assay and those kept near the ground. Similarly, minimal differences were found in the female's ability to take an additional bloodmeal after oviposition between these groups. We conclude that similar to other high-altitude migrating insects, mosquitoes are able to withstand extended high-altitude flight and subsequently reproduce and transmit pathogens by blood feeding on new hosts.

A novel fluorescence and DNA combination for versatile, long-term marking of mosquitoes.

Current mark-release-recapture methodologies are limited in their ability to address complex problems in vector biology, such as studying multiple groups overlapping in space and time. Additionally, limited mark retention, reduced post-marking survival and the large effort in marking, collection and recapture all complicate effective insect tracking.We have developed and evaluated a marking method using a fluorescent dye (SmartWater®) combined with synthetic DNA tags to informatively and efficiently mark adult mosquitoes using an airbrush pump and nebulizer. Using a handheld UV flashlight, the fluorescent marking enabled quick and simple initial detection of recaptures in a field-ready and non-destructive approach that when combined with an extraction-free PCR on individual mosquito legs provides potentially unlimited marking information.This marking, first tested in the laboratory with Anopheles gambiae s.l. mosquitoes, did not affect survival (median ages 24-28 days, p-adj > 0.25), oviposition (median eggs/female of 28.8, 32.5, 33.3 for water, green, red dyes, respectively, p-adj > 0.44) or Plasmodium competence (mean oocysts 5.56-10.6, p-adj > 0.95). DNA and fluorescence had 100% retention up to 3 weeks (longest time point tested) with high intensity, indicating marks would persist longer.We describe a novel, simple, no/low-impact and long-lasting marking method that allows separation of multiple insect subpopulations by combining unlimited length and sequence variation in the synthetic DNA tags. This method can be readily deployed in the field for marking multiple groups of mosquitoes or other insects.

Diversity, dynamics, direction, and magnitude of high-altitude migrating insects in the Sahel.

Long-distance migration of insects impacts food security, public health, and conservation-issues that are especially significant in Africa. Windborne migration is a key strategy enabling exploitation of ephemeral havens such as the Sahel, however, its knowledge remains sparse. In this first cross-season investigation (3 years) of the aerial fauna over Africa, we sampled insects flying 40-290 m above ground in Mali, using nets mounted on tethered helium-filled balloons. Nearly half a million insects were caught, representing at least 100 families from thirteen orders. Control nets confirmed that the insects were captured at altitude. Thirteen ecologically and phylogenetically diverse species were studied in detail. Migration of all species peaked during the wet season every year across localities, suggesting regular migrations. Species differed in flight altitude, seasonality, and associated weather conditions. All taxa exhibited frequent flights on southerly winds, accounting for the recolonization of the Sahel from southern source populations. "Return" southward movement occurred in most taxa. Estimates of the seasonal number of migrants per species crossing Mali at latitude 14°N were in the trillions, and the nightly distances traversed reached hundreds of kilometers. The magnitude and diversity of windborne insect migration highlight its importance and impacts on Sahelian and neighboring ecosystems.